Measuring Oxytocin and Vasopressin: Bioassays, Immunoassays and Random Numbers

May combine two claims.

The most sensitive oxytocin bioassays used mouse mammary strips and could detect 4 μU/ml (7 pg/ml) in 0.5 ml of sample, but even this was not sufficiently sensitive to measure levels in raw plasma.

“The most sensitive bioassays used mouse mammary strips and could detect 4 μU/ml (7 pg/ml) in 0.5 ml of sample 50, although even this was still not sufficiently sensitive to measure levels in raw plasma.”

• cites [50] Robinson IC, Walker JM. Bioassay of oxytocin on the superfused mammary gland of the mouse, using an automatic apparatus. Br J Pharmacol 1974; 50: 409– 417. 4368453 PMC1776677 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The human posterior pituitary contains approximately 21 μg of oxytocin, whereas that of the rat contains 0.5-1 μg.

“the human posterior pituitary contains approximately 21 μg of oxytocin 72, 73, 74, whereas that of the rat contains 0.5–1 μg.”

• cites [72] Chard T, Boyd NR, Forsling ML, McNeilly AS, Landon J. The development of a radioimmunoassay for oxytocin: the extraction of oxytocin from plasma, and its measurement during parturition in human and goat blood. J Endocrinol 1970; 48: 223– 234. 5471863 sourceNot checked yet

  Not checked in the free run.

• cites [73] Chard T, Forsling ML, James MA, Kitau MJ, Landon J. The development of a radioimmunoassay for oxytocin: sensitivity of the assay in aqueous buffer solution, specificity and the dissociation of immunological and biological activity. J Endocrinol 1970; 46: 533– 542. 5443195 sourceNot checked yet

  Not checked in the free run.

• cites [74] Chard T, Kitau MJ, Landon J. The development of a radioimmunoassay for oxytocin: radioiodination, antibody production and separation techniques. J Endocrinol 1970; 46: 269– 278. 5492003 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The normal plasma concentration of vasopressin in man is below 2 μU/ml (5 pg/ml), increasing several fold after 24 h of dehydration.

“the normal plasma concentration of vasopressin in man is below 2 μU/ml (5 pg/ml) 39, with this increasing several fold after 24 h of dehydration.”

• cites [39] Share L. Vasopressin, its bioassay and the physiological control of its release. Am J Med 1967; 42: 701– 712. 5337374 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Across mammalian species, blood volume is linearly proportional to body mass, although the mass of the pituitary is proportional only to the log of body mass.

“Across mammalian species, blood volume is linearly proportional to body mass, although the mass of the pituitary is proportional only to the log of body mass 71.”

• cites [71] Stahl WR. Organ weights in primates and other mammals. Science 1965; 150: 1039– 1042. 5843618 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

In lactating rats, each suckling-induced burst of oxytocin neurones is followed by an increase in intramammary pressure that can be mimicked by i.v. injection of approximately 0.5 mU (0.8 ng) of oxytocin.

“Each burst is followed by an increase in intramammary pressure that can be mimicked by i.v. injection of approximately 0.5 mU (0.8 ng) of oxytocin 16, 65.”

• cites [16] Leng G, Pineda R, Sabatier N, Ludwig M. 60 YEARS OF NEUROENDOCRINOLOGY: the posterior pituitary, from Geoffrey Harris to our present understanding. J Endocrinol 2015; 226: T173– T185. 25901040 sourceNot checked yet

  Not checked in the free run.

• cites [65] Wakerley JB, Lincoln DW. Intermittent release of oxytocin during suckling in the rat. Nat New Biol 1971; 233: 180– 181. 5287087 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The kidney responds to changes in the rate of vasopressin secretion of less than 1 pg/min/kg, and the rate required to produce isosmolar urine during overhydration is approximately 3 pg/min/kg.

“the kidney responds to changes in the rate of vasopressin secretion of < 1 pg/min/kg, and also that the rate required to produce isosmolar urine during overhydration is approximately 3 pg/min/kg”

• cites [44] Andersen LJ, Andersen JL, Schutten HJ, Warberg J, Bie P. Antidiuretic effect of subnormal levels of arginine vasopressin in normal humans. Am J Physiol 1990; 259: R53– R60. 2375429Not checked yet

  Not checked in the free run.

• cites [45] Hollander W Jr, Williams TF, Fordham CC 3rd, Welt LG. A study of the quantitative relationship between antidiuretic hormone (vasopressin) and the renal tubular reabsorption of water. J Clin Invest 1957; 36: 1059– 1071. 13449158 PMC1072692 sourceNot checked yet

  Not checked in the free run.

• cites [46] Raisz LG, Mc NW, Saxon L. Studies on the renal concentrating mechanism. I. Role of vasopressin. J Lab Clin Med 1958; 52: 437– 445. 13575934Not checked yet

  Not checked in the free run.

May combine two claims.

Saameli calculated that the half-life of oxytocin in pregnant women was approximately 3-4 min, and that uterine contractions were initiated by infusions that raised plasma concentrations by 3 μU/ml (5 pg/ml).

“he calculated that the half‐life of oxytocin in pregnant women was approximately 3–4 min, and also that uterine contractions were initiated by infusions that raised plasma concentrations by 3 μU/ml (5 pg/ml).”

• cites [52] Saameli K. An indirect method for the estimation of oxytocin blood concentration and half‐life in pregnant women near term. Am J Obstet Gynecol 1963; 85: 186– 192. 13975829 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The plasma volume of a rat is approximately 4 ml/100 g, and a burst that releases 0.5 mU (0.8 ng) oxytocin should raise circulating concentrations in a 300 g rat by approximately 70 pg/ml.

“The plasma volume of a rat is approximately 4 ml/100 g 67, and so a burst that releases 0.5 mU (0.8 ng) oxytocin should raise circulating concentrations in a 300 g rat by approximately 70 pg/ml.”

• cites [67] Lee HB, Blaufox MD. Blood volume in the rat. J Nucl Med 1985; 26: 72– 76. 3965655Not checked yet

  Not checked in the free run.

May combine two claims.

Higuchi et al. developed a radioimmunoassay that allowed oxytocin to be measured in unextracted rat plasma, and during suckling-induced milk let-down plasma concentrations rose from 16.5 pg/ml to approximately 65 pg/ml, declining with a half-life of approximately 1.5 min.

“Higuchi et al. 68 developed a radioimmunoassay that allowed oxytocin to be measured in unextracted rat plasma, and took samples at the time of suckling‐induced milk let‐down, as recognised by stretching movements of the suckling pups. Plasma concentrations rose from 16.5 pg/ml to approximately 65 pg/ml, and declined with a half‐life of approximately 1.5 min.”

• cites [68] Higuchi T, Tadokoro Y, Honda K, Negoro H. Detailed analysis of blood oxytocin levels during suckling and parturition in the rat. J Endocrinol 1986; 110: 251– 256. 3755747 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

In pigs, suckling evokes increases in intramammary pressure equivalent to those induced by i.v. injection of 50 mU (83 ng) of oxytocin, which for a plasma volume of 5 l would raise plasma concentrations by approximately 17 pg/ml, close to the peak concentration (approximately 18 pg/ml) measured by radioimmunoassay during suckling-induced milk ejection.

“In pigs (body weight 150–200 kg), suckling evokes increases in intramammary pressure equivalent to those induced by i.v. injection of 50 mU (83 ng) of oxytocin 69. For a plasma volume of 5 l, these would raise plasma concentrations by approximately 17 pg/ml, which is close to the peak concentration (approximately 18 pg/ml) measured by radioimmunoassay during suckling‐induced milk ejection.”

• cites [69] Ellendorff F, Forsling ML, Poulain DA. The milk ejection reflex in the pig. J Physiol 1982; 333: 577– 594. 6892113 PMC1197265 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Such interference has been recognised in immunoassays for cholecystokinin, parathyroid hormone, thyrotropic hormone, glucagon, secretin, angiotensin, growth hormone, vasoactive intestinal peptide, gonadotrophins and gonadotrophin-releasing hormone, all of which were resolved by sample extraction.

“Such interference has also been recognised in immunoassays for cholecystokinin 80, parathyroid hormone 81, thyrotropic hormone 82, glucagon 83, secretin 84, 85, angiotensin 86, growth hormone 87, vasoactive intestinal peptide 88, gonadotrophins 89 and gonadotrophin‐releasing hormone 90, all of which were resolved by sample extraction.”

• cites [80] Harvey RF, Dowsett L, Hartog M, Read AE. Radioimmunoassay of cholecystokinin‐pancreozymin. Gut 1974; 15: 690– 699. 4435583 PMC1413038 sourceNot checked yet

  Not checked in the free run.

• cites [81] Christensen MS. A sensitive radioimmunoassay of parathyroid hormone in human serum using a specific extraction procedure. Scand J Clin Lab Invest 1976; 36: 313– 322. 959749 sourceNot checked yet

  Not checked in the free run.

• cites [82] Spencer CA, Challand GS. Interference in a radioimmunoassay for human thyrotropin. Clin Chem 1977; 23: 584– 588. 556979 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The first peak accounted for a 67% loss of binding of antibody to vasopressin, and the second peak accounted for a further 30% loss; neither apparent immunoreactivity was displaced by excess cold vasopressin, indicating occlusion of antibody recognition rather than competitive binding.

“The first peak accounted for a 67% loss of binding of antibody to vasopressin, and the second peak accounted for a further 30% loss. Neither of these apparent ‘immunoreactivities’ were displaced by an excess of cold vasopressin, indicating that they did not involve competitive binding of vasopressin, but occlusion of antibody recognition of vasopressin.”

• cites [78] Robertson GL, Klein LA, Roth J, Gorden P. Immunoassay of plasma vasopressin in man. Proc Natl Acad Sci USA 1970; 66: 1298– 1305. 5273455 PMC335819 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

It is unlikely that an antibody will recognise a different ligand with higher affinity than it has for the ligand to which it was raised; it may recognise other ligands with lower affinity, although this presents a major problem only if that different ligand is present at higher concentrations.

“However, it is unlikely that an antibody will recognise a different ligand with higher affinity than it has for the ligand to which it was raised; it may recognise other ligands with lower affinity, although this presents a major problem only if that different ligand is present at higher concentrations.”

Classified as probably not a substantial claim.

Close attention was given to extraction and concentration procedures in developing immunoassays for vasopressin and oxytocin.

“Thus, close attention was given to extraction and concentration procedures 76.”

• cites [76] Chard T. The radioimmunoassay of oxytocin and vasopressin. J Endocrinol 1973; 58: 143– 160. 4578963 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

In unextracted plasma from healthy subjects, Fyhrquist et al. found a mean vasopressin concentration of 2 pg/ml supine and 6.2 pg/ml upright, in agreement with bioassay levels measured by Segar and Moore.

“In unextracted plasma from healthy subjects, they found a mean vasopressin concentration of 2 pg/ml in the supine position and 6.2 pg/ml in the upright position, which is in agreement with the levels measured in bioassays by Segar and Moore 33.”

• cites Fyhrquist et al. 92Not checked yet

  Not checked in the free run.

• cites Segar and Moore 33Not checked yet

  Not checked in the free run.

May combine two claims.

Robertson et al. resolved three peaks of immunoreactivity by filtering plasma on G-25 Sephadex; only the third and smallest was consistent with true vasopressin, being recovered in the same location as pure vasopressin.

“By filtering plasma samples on G‐25 Sephadex, they resolved three peaks of ‘immunoreactivity’, only the third and smallest of which was consistent with true vasopressin, being recovered in the same location as pure vasopressin 78.”

• cites [78] Robertson GL, Klein LA, Roth J, Gorden P. Immunoassay of plasma vasopressin in man. Proc Natl Acad Sci USA 1970; 66: 1298– 1305. 5273455 PMC335819 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Enzymatic degradation of oxytocin is a problem only when measuring in plasma from pregnant women, and extraction procedures can remove such enzymes efficiently.

“Fortunately, enzymatic degradation of oxytocin is a problem only when measuring in plasma from pregnant women 54, and extraction procedures can remove such enzymes efficiently.”

• cites [54] Leake RD, Weitzman RE, Glatz TH, Fisher DA. Plasma oxytocin concentrations in men, nonpregnant women, and pregnant women before and during spontaneous labor. J Clin Endocrinol Metab 1981; 53: 730– 733. 7287862 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

In 1973, Chard noted that the best immunoassays for vasopressin were no more sensitive than the best bioassays; the same is still true today for most radioimmunoassays and most enzyme-linked immunoassays (ELISAs).

“In 1973, Chard 76 noted that best immunoassays for vasopressin were no more sensitive than the best bioassays; the same is still true today for most radioimmunoassays and most enzyme‐linked immunoassays (ELISAs).”

• cites [76] Chard T. The radioimmunoassay of oxytocin and vasopressin. J Endocrinol 1973; 58: 143– 160. 4578963 sourceNot checked yet

  Not checked in the free run.

May combine two claims. The quote only partly states this claim.

The third peak reproduced the expected relationship with plasma osmotic pressure in normal adults, the expected high levels in nephrogenic diabetes insipidus, and the expected low levels in pituitary diabetes insipidus.

“The immunoreactivity in the third peak reproduced the expected relationship with plasma osmotic pressure in normal adults, the expected high levels in nephrogenic diabetes insipidus”

• cites [78] Robertson GL, Klein LA, Roth J, Gorden P. Immunoassay of plasma vasopressin in man. Proc Natl Acad Sci USA 1970; 66: 1298– 1305. 5273455 PMC335819 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

After accounting for plasma matrix interference, Robertson et al. calculated that normal plasma concentrations of vasopressin in man were approximately 1 pg/ml, increasing to approximately 10 pg/ml with dehydration.

“After accounting for the plasma matrix interference, Robertson et al. 78 calculated that normal plasma concentrations of vasopressin in man were approximately 1 pg/ml, increasing to approximately 10 pg/ml with dehydration.”

• cites [78] Robertson GL, Klein LA, Roth J, Gorden P. Immunoassay of plasma vasopressin in man. Proc Natl Acad Sci USA 1970; 66: 1298– 1305. 5273455 PMC335819 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

There were considerable difficulties in developing immunoassays for vasopressin and oxytocin because they have a low molecular weight, are poor immunogens, making high-affinity antisera difficult, and circulating levels were clearly very low.

“There were considerable difficulties in developing immunoassays for vasopressin and oxytocin. Because these have a low molecular weight, they are poor immunogens, making the preparation of high‐affinity antisera difficult, and circulating levels were clearly very low.”

• cites [76] Chard T. The radioimmunoassay of oxytocin and vasopressin. J Endocrinol 1973; 58: 143– 160. 4578963 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

By the end of the 20th century, bioassays were still of critical importance in the discovery of novel antagonists and agonists of vasopressin and oxytocin because biological activity cannot be inferred from structure alone.

“By the end of the 20th century, bioassays were still ‘of critical importance’ in the discovery of novel antagonists and antagonists of vasopressin and oxytocin because biological activity cannot be inferred from structure alone 75.”

• cites [75] Chan WY, Wo NC, Stoev ST, Cheng LL, Manning M. Discovery and design of novel and selective vasopressin and oxytocin agonists and antagonists: the role of bioassays. Exp Physiol 2000; 85S: 7S– 18S. 10795902 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Overestimation can arise when enzymes are present that degrade oxytocin; the plasma of late pregnant women contains abundant oxytocinase, which inactivates oxytocin by splitting the peptide linkage between the cysteine and tyrosine residues.

“Overestimation can also arise when enzymes are present that degrade oxytocin. The plasma of late pregnant women contains abundant oxytocinase, which inactivates oxytocin by splitting the peptide linkage between the cysteine and tyrosine residues 102.”

• cites [102] Nomura S, Ito T, Yamamoto E, Sumigama S, Iwase A, Okada M, Shibata K, Ando H, Ino K, Kikkawa F, Mizutani S. Gene regulation and physiological function of placental leucine aminopeptidase/oxytocinase during pregnancy. Biochim Biophys Acta 2005; 1751: 19– 25. 15894523 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

By the 1980s, many laboratories had generated radioimmunoassays for oxytocin using independently produced antibodies, and most but not all required extraction.

“By the 1980s, many laboratories had generated radioimmunoassays for oxytocin, using independently produced antibodies 58, 72, 73, 74, 94, 95, 96, 97, 98] and most, but not all 99, 100, required extraction.”

• cites [58] Dawood MY. Evolving concepts of oxytocin for induction of labor. Am J Perinatol 1989; 6: 167– 172. 2653335 sourceNot checked yet

  Not checked in the free run.

• cites [72] Chard T, Boyd NR, Forsling ML, McNeilly AS, Landon J. The development of a radioimmunoassay for oxytocin: the extraction of oxytocin from plasma, and its measurement during parturition in human and goat blood. J Endocrinol 1970; 48: 223– 234. 5471863 sourceNot checked yet

  Not checked in the free run.

• cites [73] Chard T, Forsling ML, James MA, Kitau MJ, Landon J. The development of a radioimmunoassay for oxytocin: sensitivity of the assay in aqueous buffer solution, specificity and the dissociation of immunological and biological activity. J Endocrinol 1970; 46: 533– 542. 5443195 sourceNot checked yet

  Not checked in the free run.

• cites [74] Chard T, Kitau MJ, Landon J. The development of a radioimmunoassay for oxytocin: radioiodination, antibody production and separation techniques. J Endocrinol 1970; 46: 269– 278. 5492003 sourceNot checked yet

  Not checked in the free run.

• cites [94] Dawood MY, Raghavan KS, Pociask C. Radioimmunoassay of oxytocin. J Endocrinol 1978; 76: 261– 270. 627820 sourceNot checked yet

  Not checked in the free run.

• cites [95] Schams D. Oxytocin determination by radioimmunoassay. III. Improvement to subpicogram sensitivity and application to blood levels in cyclic cattle. Acta Endocrinol (Copenh) 1983; 103: 180– 183. 6683055Not checked yet

  Not checked in the free run.

• cites [96] Chiodera P, Louis F, Legros JJ. Simultaneous radioimmunoassay for plasma arginine‐vasopressin and oxytocin using DEAE Sephadex A 25 extraction. J Endocrinol Invest 1984; 7: 287– 293. 6501802 sourceNot checked yet

  Not checked in the free run.

• cites [97] Sheldrick EL, Flint AP. Circulating concentrations of oxytocin during the estrous cycle and early pregnancy in sheep. Prostaglandins 1981; 22: 631– 636. 7323312 sourceNot checked yet

  Not checked in the free run.

• cites [98] Landgraf R. Plasma oxytocin concentrations in man after different routes of administration of synthetic oxytocin. Exp Clin Endocrinol 1985; 85: 245– 248. 4018162 sourceNot checked yet

  Not checked in the free run.

• cites [99] Burd JM, Weightman DR, Baylis PH. Solid phase radioimmunoassay for direct measurement of human plasma oxytocin. J Immunoassay 1985; 6: 227– 243. 3900138 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

When oxytocin is incubated in late-pregnancy plasma, there is a rapid loss of biological activity but after 2 h oxytocin immunoreactivity exceeds bioactivity by three-fold; the explanation is that oxytocinase degrades labelled oxytocin, reducing label binding to antibody and leading to overestimation.

“When oxytocin is incubated in this plasma, there is a rapid loss of biological activity but, after 2 h, oxytocin ‘immunoreactivity’ exceeds bioactivity by three‐fold. The obvious explanation is that oxytocinase in the sample also degrades the labelled oxytocin. The consequence will be that less label will bind to antibody, leading to overestimation of the amount of oxytocin 76.”

• cites [76] Chard T. The radioimmunoassay of oxytocin and vasopressin. J Endocrinol 1973; 58: 143– 160. 4578963 sourceNot checked yet

  Not checked in the free run.

• AssumptionRests on an assumption: When oxytocin or vasopressin are secreted, the entire contents of the vesicle are discharged, including elements of incompletely processed precursor that may not be biologically active.

May combine two claims.

In 1970, Robertson et al. described one of the first radioimmunoassays to measure plasma vasopressin in man; it was sensitive to 1 pg/ml (0.3 mU/ml) but when whole plasma was assayed, apparent concentrations were several hundredfold higher than anticipated, failed to fluctuate appropriately in response to physiological manipulation or disease states, and were not reduced substantially by dialysis.

“This assay was sensitive to 1 pg/ml (0.3 mU/ml) but, when whole plasma was assayed, apparent concentrations were ‘several hundredfold higher than anticipated, failed to fluctuate appropriately in response to physiological manipulation or in disease states, and were not reduced substantially by dialysis of the plasma under conditions that completely removed vasopressin’.”

• cites [78] Robertson GL, Klein LA, Roth J, Gorden P. Immunoassay of plasma vasopressin in man. Proc Natl Acad Sci USA 1970; 66: 1298– 1305. 5273455 PMC335819 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

An antibody might recognise elements unrelated to the antigen against which it was raised, and unrelated molecules might cross-react.

“In some cases, an antibody might recognise elements unrelated to the antigen against which it was raised. Antibodies are sensitive to differences in amino acid sequence, although they recognise the structure of a molecule, not its amino acid sequence per se, and so unrelated molecules might cross‐react 76, 79.”

• cites [76] Chard T. The radioimmunoassay of oxytocin and vasopressin. J Endocrinol 1973; 58: 143– 160. 4578963 sourceNot checked yet

  Not checked in the free run.

• cites [79] Martin MJ, Chard T, Landon J. The development of a radioimmunoassay for bovine neurophysin. J Endocrinol 1972; 52: 481– 495. 4336118 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

At that time, seven other vasopressin radioimmunoassays, using different antibodies and extraction procedures, all reported plasma concentrations of 1-10 pg/ml, and all showed that concentrations changed with physiological conditions in the manner expected from bioassays.

“At that time, seven other vasopressin radioimmunoassays had been described, using different antibodies and different extraction procedures. All had reported plasma concentrations of 1–10 pg/ml, and all showed that concentrations changed with physiological conditions in the manner expected from bioassays.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Baylis and Heath emphasised that even small increases in protein concentration lower binding and careful control of extract protein concentration is needed to avoid misinterpretation that a fall in binding is due to immunoreactive vasopressin.

“When describing their vasopressin assay, Baylis and Heath 91 emphasised that ‘even small increases in protein concentration lower binding and therefore careful control of extract protein concentration is needed to avoid the misinterpretation that a fall in binding is due to immunoreactive vasopressin’.”

• cites [91] Baylis PH, Heath DA. The development of a radioimmunoassay for the measurement of human plasma arginine vasopressin. Clin Endocrinol (Oxf) 1977; 7: 91– 102. 891002 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Antibodies do not recognise the whole molecule, so an immunoassay may overestimate hormone concentrations by measuring fragments of no physiological significance.

“Antibodies do not recognise the whole molecule, and so an immunoassay may overestimate hormone concentrations by measuring fragments of no physiological significance 76.”

• cites [76] Chard T. The radioimmunoassay of oxytocin and vasopressin. J Endocrinol 1973; 58: 143– 160. 4578963 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

With few exceptions, assays in extracted plasma reported basal oxytocin levels below 5 pg/ml in man; of 17 studies reviewed by Szeto et al., only two reported higher levels, the highest being 71 pg/ml in samples from the corpus cavernosum of the penis.

“With few exceptions, assays in extracted plasma reported basal levels of < 5 pg/ml in man: of 17 studies reviewed by Szeto et al. 101, only two reported higher levels; the highest (71 pg/ml) were in samples taken from the corpus cavernosum of the penis, a circumstance that might be expected to affect hormone secretion.”

• cites Szeto et al. 101Not checked yet

  Not checked in the free run.

May combine two claims.

Henneberry et al. assayed unextracted samples using standard curves generated in charcoal-filtered plasma and reported basal vasopressin levels of 1.8 pg/ml, rising to 6.3 pg/ml after a saline infusion that raised plasma osmotic pressure from 280 to 293 mOsm/l.

“Henneberry et al. 93 took a different approach: they assayed unextracted samples using standard curves generated in charcoal‐filtered plasma, and reported basal levels of 1.8 pg/ml, rising to 6.3 pg/ml after a saline infusion that raised plasma osmotic pressure from 280 to 293 mOsm/l.”

• cites Henneberry et al. 93Not checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

High salt concentrations have a nonlinear effect, and serial dilutions of samples with a high salt content produce measurements that do not parallel the standard curve.

“High salt concentrations have a nonlinear effect, and serial dilutions of samples with a high salt content produce measurements that do not parallel the standard curve.”

May combine two claims.

To sustain a plasma concentration of 350 pg/ml in man, oxytocin must be produced at 40,000 times the rate measured in the rat; given a difference in pituitary mass and oxytocin content of just 40-fold, this seems beyond plausibility.

“Thus, to sustain a plasma concentration of 350 pg/ml in man, oxytocin must be produced at 40 000 times the rate measured in the rat. Given a difference in pituitary mass and oxytocin content of just 40‐fold, this seems beyond plausibility.”

May combine two claims.

In rats, Ginsburg and Heller estimated that the initial half-life of exogenously administered vasopressin was less than 1 min; from experiments in acutely nephrectomised rats and by ligating arteries to reduce liver blood flow, they concluded that most vasopressin was cleared by the kidneys and liver.

“In rats, Ginsburg and Heller 104 estimated that the initial half‐life of exogenously administered vasopressin was less than 1 min. From experiments in acutely nephrectomised rats, and by ligating arteries to reduce blood flow through the liver, it was concluded that most of the vasopressin was cleared by the kidneys and liver.”

• cites Ginsburg and Heller 104Not checked yet

  Not checked in the free run.

May combine two claims.

Claims that the Enzo ELISA is a valid way of measuring oxytocin in unextracted plasma do not rest on the manufacturer's advice, which strongly recommends extraction, but instead converge on two papers, neither of which actually investigated human plasma.

“Claims that the Enzo ELISA is a valid way of measuring oxytocin in unextracted plasma do not rest on manufacturer's advice, which, as we have noted, strongly recommends extraction, but, instead, converge on two papers, neither of which actually investigated human plasma at all.”

May combine two claims.

Ginsburg and Smith found that oxytocin added to rat plasma in vitro at 37 °C was relatively stable, losing just 6-13% of biological activity after 40 min, whereas when oxytocin was injected into rats its biological activity disappeared with a half-life of 1.65 min.

“Ginsburg and Smith 49, using a rat uterus bioassay, found that, when oxytocin was added to rat plasma in vitro at 37 °C, it was relatively stable, losing just 6–13% of biological activity after 40 min of incubation. By contrast, when oxytocin was injected into rats, the biological activity disappeared with a half‐life of 1.65 min.”

• cites Ginsburg and Smith 49Not checked yet

  Not checked in the free run.

May combine two claims.

In dogs, Ang and Jenkins injected radiolabelled vasopressin and used thin-layer chromatography and autoradiography to quantify labelled vasopressin in plasma; it disappeared with an initial half-life of 3 min and a later half-life of 13 min.

“In dogs, rather than measure vasopressin by either radioimmunoassay or bioassay, Ang and Jenkins 106 injected radiolabelled vasopressin and used thin‐layer chromatography and autoradiography to quantify labelled vasopressin in plasma samples: it disappeared with an initial half‐life of 3 min and a later half‐life of 13 min.”

• cites Ang and Jenkins 106Not checked yet

  Not checked in the free run.

May combine two claims.

In patients with chronic renal failure, vasopressin clearance is much slower, with metabolic clearance rate reduced by approximately half, consistent with the conclusion that vasopressin is mainly cleared from the circulation through the kidneys.

“In patients with chronic renal failure, the clearance is much slower: MCR is reduced by approximately half 108, which is consistent with the conclusion that vasopressin is mainly cleared from the circulation through the kidneys.”

• cites [108] Argent NB, Wilkinson R, Baylis PH. Metabolic clearance rate of arginine vasopressin in severe chronic renal failure. Clin Sci (Lond) 1992; 83: 583– 587. 1335395 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Ultrafiltration of plasma indicated that approximately 30% of similarly infused vasopressin was bound to large molecules, whereas there was no apparent binding of oxytocin; in vitro oxytocin bioactivity was stable, with only approximately 20% lost after 24 h at 37 °C.

“Ultrafiltration of the plasma indicated that, whereas approximately 30% of similarly infused vasopressin was bound to large molecules, there was no apparent binding of oxytocin. In vitro, oxytocin bioactivity was stable, as found in rats 49; only approximately 20% was lost after 24 h at 37 °C.”

• cites [49] Ginsburg M, Smith MW. The fate of oxytocin in male and female rats. Br J Pharmacol Chemother 1959; 14: 327– 333. 13850241 PMC1481739 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The reported mean oxytocin and vasopressin concentrations of typically 100-500 pg/ml in unextracted human plasma are inconsistent by two orders of magnitude with bioassays and with radioimmunoassay measurements on extracted samples, and are also inconsistent with the known sensitivity of peripheral targets and with what is known of the production, storage and clearance of these hormones.

“These values are inconsistent, by two orders of magnitude, with both bioassays of oxytocin and vasopressin and with radioimmunoassay measurements on extracted samples. They are also inconsistent with the known sensitivity of peripheral targets to oxytocin and vasopressin, and with what is known of the production, storage and clearance of these hormones.”

May combine two claims.

In nephrectomised rats, oxytocin bioactivity declined with a half-life of approximately 3 min in the first 7 min and much more slowly thereafter; when the splanchnic vessels were also ligated, the initial decline was not further changed but after 7 min oxytocin concentrations stuck at a high, steady level.

“In nephrectomised rats, bioactivity declined with a half‐life of approximately 3 min in the first 7 min, and much more slowly thereafter. When the splanchnic vessels were also ligated, the initial decline was not further changed but, after 7 min, oxytocin concentrations ‘stuck’ at a high, steady level.”

• cites Ginsburg and Smith 49Not checked yet

  Not checked in the free run.

May combine two claims.

Ginsburg and Smith deduced that when oxytocin enters the circulation it is rapidly removed by the kidneys and liver, and that without these organs injected oxytocin diffuses within approximately 7 min throughout the extracellular fluid, where it is degraded only very slowly.

“Ginsburg and Smith deduced that, when oxytocin enters the circulation, it is rapidly removed by the kidneys and liver. Without these organs, injected oxytocin diffuses within approximately 7 min throughout the extracellular fluid, where it is degraded only very slowly.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Zhang et al. recognised that one common challenge in biomarker analysis is to select an appropriate surrogate matrix to prepare the standard curve, free of target analytes and identical or similar to unknown sample matrix with regard to ionisation effects and extraction recovery.

“As Zhang et al. 125 recognised, ‘One common challenge in biomarker analysis is to select an appropriate surrogate matrix to prepare the standard curve, which is free of target analytes and is identical or similar to unknown sample matrix with regard to ionisation effects and extraction recovery’.”

• cites Zhang et al. 125Not checked yet

  Not checked in the free run.

May combine two claims.

In man, Fabian et al. measured oxytocin with a mammary gland bioassay and found that after the end of an i.v. infusion bioactivity disappeared with a half-life of 4.8 min and a distribution volume of 10.5 ml/100 g, close to the extracellular fluid volume, whereas an acute large injection disappeared with a half-life of 3.2 min and a distribution volume of 7.4 ml/100 g.

“In man, Fabian et al. 43 measured oxytocin with a mammary gland bioassay and studied its clearance after the end of an i.v. infusion. Bioactivity disappeared with a half‐life of 4.8 min with a distribution volume of 10.5 ml/100 g, which is close to the extracellular fluid volume, whereas an acute large injection disappeared with a half‐life of 3.2 min and a distribution volume of 7.4 ml/100 g.”

• cites Fabian et al. 43Not checked yet

  Not checked in the free run.

May combine two claims. The quote only partly states this claim.

Three studies compared measurements of oxytocin in the same samples by the same ELISA with and without extraction; Robinson et al. found approximately 8 pg/ml in extracted seal plasma versus 300 pg/ml in unextracted heparin samples or 540 pg/ml in unextracted EDTA samples with no correlation, Christensen et al. found radioimmunoassay extracted mean levels of 1.1 pg/ml and spiked levels higher by t

“Three studies have compared measurements of oxytocin in the same samples by the same ELISA, with and without extraction 101, 116, 117. Robinson et al. 116 measured oxytocin levels in seals: in extracted samples, they found plasma levels of approximately 8 pg/ml, compared to 300 pg/ml in unextracted samples collected in heparin vacutainers or 540 pg/ml in unextracted samples collected in ethylenedi”

• cites [101] Szeto A, McCabe PM, Nation DA, Tabak BA, Rossetti MA, McCullough ME, Schneiderman N, Mendez AJ. Evaluation of enzyme immunoassay and radioimmunoassay methods for the measurement of plasma oxytocin. Psychosom Med 2011; 73: 393– 400102. 21636661 PMC3118424 sourceNot checked yet

  Not checked in the free run.

• cites [116] Robinson KJ, Hazon N, Lonergan M, Pomeroy PP. Validation of an enzyme‐linked immunoassay (ELISA) for plasma oxytocin in a novel mammal species reveals potential errors induced by sampling procedure. J Neurosci Meth 2014; 226: 73– 79. 24485867 sourceNot checked yet

  Not checked in the free run.

• cites [117] Christensen JC, Shiyanov PA, Estepp JR, Schlager JJ. Lack of association between human plasma oxytocin and interpersonal trust in a Prisoner's Dilemma paradigm. PLoS ONE 2014; 9: e116172. 25549255 PMC4280178 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

As a proportion of body weight, the rat pituitary contains much more oxytocin and vasopressin than the human pituitary, and in rats and other small mammals circulating levels are higher; in man and other large animals lower circulating concentrations are compensated for by higher sensitivity of peripheral targets.

“As a proportion of body weight, the rat pituitary contains much more oxytocin and vasopressin than the human pituitary and, in rats, as in other small mammals, circulating levels are higher. In man and other large animals, lower circulating concentrations are compensated for by higher sensitivity of the peripheral targets.”

May combine two claims.

In rabbits, vasopressin was cleared with a half-life of approximately 4.5 min, mainly by the kidneys and liver.

“In similar studies on rabbits, vasopressin was cleared with a half‐life of approximately 4.5 min, again mainly by the kidneys and liver 105.”

• cites [105] Hasan SH. The clearance of vasopressin and deaminovasopressin from the circulation of normal and hypercalcaemic rabbits. J Endocrinol 1970; 47: 135– 136. 5428912 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Plasma oxytocin was measured using human plasma diluted with water (1:6) as a surrogate matrix, and basal levels of 2.4 pg/ml for human but 298 pg/ml for rat were reported; these high levels in the rat may indicate that human plasma is not a suitable surrogate matrix for measurements in rat.

“Plasma oxytocin was measured using human plasma diluted with water (1 : 6) as a surrogate matrix, and basal levels of 2.4 pg/ml for human but 298 pg/ml for rat were reported; these high levels in the rat may indicate that human plasma is not a suitable surrogate matrix for measurements in rat.”

• cites [125] Zhang G, Zhang Y, Fast DM, Lin Z, Steenwyk R. Ultra sensitive quantitation of endogenous oxytocin in rat and human plasma using a two‐dimensional liquid chromatography‐tandem mass spectrometry assay. Anal Biochem 2011; 416: 45– 52. 21609710 sourceNot checked yet

  Not checked in the free run.

Classified as probably not a substantial claim. May combine two claims.

The Enzo ELISAs draw attention to plasma matrix interference, illustrate the benefits of plasma extraction, and strongly recommend that users extract plasma samples; they state that dilution might minimise interference but warn that users must verify that dilutions are appropriate for their samples.

“The ELISAs in most widespread use are marketed by Enzo, who draw attention to problems of plasma matrix interference, illustrate the benefits of plasma extraction, and ‘strongly recommend’ that users extract plasma samples. They state that it might be possible to dilute plasma samples enough to minimise interference (suggesting at least an eight‐fold dilution) but warn that the user ‘must verify’”

• cites [113] Designs A. Oxytocin Enzyme Immunoassay (EIA) kit: Manufacturer's instructions. Available at: http://www.enzolifesciences.com/fileadmin/redacteur/pdf/adi/ADI-900-153.pdf.Not checked yet

  Not checked in the free run.

May combine two claims.

Carter et al. reported that in voles the ELISA measured spiked levels of oxytocin with a gain of 0.79, consistent with a reduction in antibody affinity although not sufficient to account for the discrepancy; in HPLC fractions apparent immunoreactivity appeared only in fractions that eluted with oxytocin, suggesting no plasma interference, but the same method had previously been used by Wismer

“Carter et al. 119 further reported that, in voles, the ELISA measured spiked levels of oxytocin with a gain of 0.79. This is consistent with a reduction in antibody affinity, although not sufficient to account for the discrepancy between expected and measured oxytocin in Kramer's measurements. In fractions separated by high‐performance liquid chromatography (HPLC), apparent immunoreactivity”

• cites Carter et al. 119Not checked yet

  Not checked in the free run.

• cites Wismer Fries et al. 120Not checked yet

  Not checked in the free run.

• cites Anderson 121Not checked yet

  Not checked in the free run.

May combine two claims.

Lauson and Bacanegra concluded that at physiological levels below 25 pg/ml most plasma vasopressin is bound to protein and cleared by tubular cells of the kidney by enzymatic inactivation and secretion, while at higher concentrations more vasopressin is unbound and some is filtered and excreted into urine.

“Lauson and Bacanegra 103 concluded that, at physiological levels (< 25 pg/ml), most of the vasopressin in the plasma is bound to protein and is cleared by the tubular cells of the kidney by enzymatic inactivation and secretion. At higher concentrations, more of the vasopressin is unbound, and some of this is filtered at the glomerulus and excreted into urine.”

• cites Lauson and Bacanegra 103Not checked yet

  Not checked in the free run.

May combine two claims.

Binding of plasma proteins to an immunoglobulin can hinder its ability to bind an antigen, reducing its affinity, and/or may completely prevent some of the antibody from binding to antigen.

“The binding of plasma proteins to an immunoglobulin can hinder its ability to bind an antigen, reducing its affinity, and/or may completely prevent some of the antibody from binding to antigen.”

• cites Bjerner et al. 111Not checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

A plasma concentration of 350 pg/ml oxytocin in man implies a total blood content of approximately 1 μg given a plasma volume of 3 l, and for a half-life of 10 min implies a secretion and production rate of 500 ng/min, or 720 IU/day.

“A plasma concentration of 350 pg/ml oxytocin in man implies a total blood content of approximately 1 μg (given a plasma volume of 3 l) and, for a half‐life of 10 min, this implies a secretion (and production) rate of 500 ng/min, or 720 IU/day.”

May combine two claims.

In the baboon, oxytocin clearance was consistent with a two-compartmental system corresponding to the plasma volume and the extravascular fluid volume; the initial half-life in the first phase of distribution was approximately 1 min, followed by a terminal elimination half-life of approximately 10 min.

“In the baboon, oxytocin clearance was also consistent with a two‐compartmental system, corresponding to the plasma volume and the extravascular fluid volume. The initial half‐life in the first phase of distribution was approximately 1 min, followed by a terminal elimination half‐life of approximately 10 min 109.”

• cites [109] Kowalski WB, Diveky L, Mehendale R, Parsons M, Wilson L Jr. Effect of pregnancy on the metabolic clearance rate and the volume of distribution of oxytocin in the baboon. Am J Physiol 1998; 274: E791– E795. 9612235 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The manufacturer's advice to extract plasma has been largely ignored; many studies have used the Enzo assay on unextracted human plasma, reporting mean concentrations of oxytocin and vasopressin typically 100-500 pg/ml, and those that diluted samples did so by less than the manufacturer's recommendations without confirming that their dilution was adequate.

“This advice has been largely ignored. Many studies have used this assay on unextracted human plasma, reporting mean concentrations of oxytocin and vasopressin of typically 100–500 pg/ml 3, 4, 5, 6, 114. Those that have diluted samples have done so by less than the manufacturer's recommendations, without confirming that their dilution is adequate.”

• cites [3] Jobst A, Dehning S, Ruf S, Notz T, Buchheim A, Henning‐Fast K, Meissner D, Meyer S, Bondy B, Muller N, Zill P. Oxytocin and vasopressin levels are decreased in the plasma of male schizophrenia patients. Acta Neuropsychiatr 2014; 26: 347– 355. 25288094 sourceNot checked yet

  Not checked in the free run.

• cites [4] Apter‐Levi Y, Zagoory‐Sharon O, Feldman R. Oxytocin and vasopressin support distinct configurations of social synchrony. Brain Res 2014; 1580: 124– 132. 24184577 sourceNot checked yet

  Not checked in the free run.

• cites [5] Gouin JP, Pournajafi‐Nazarloo H, Carter CS. Changes in social functioning and circulating oxytocin and vasopressin following the migration to a new country. Physiol Behav 2015; 139: 67– 72. 25446216 sourceNot checked yet

  Not checked in the free run.

• cites [6] Rubin LH, Carter CS, Bishop JR, Pournajafi‐Nazarloo H, Drogos LL, Hill SK, Ruocco AC, Keedy SK, Reilly JL, Keshavan MS, Pearlson GD, Tamminga CA, Gershon ES, Sweeney JA. Reduced levels of vasopressin and reduced behavioral modulation of oxytocin in psychotic disorders. Schizophr Bull 2014; 40: 1374– 1384. 24619535 PMC4193712 sourceNot checked yet

  Not checked in the free run.

• cites [114] McCullough ME, Churchland PS, Mendez AJ. Problems with measuring peripheral oxytocin: can the data on oxytocin and human behavior be trusted? Neurosci Biobehav Rev 2013; 37: 1485– 1492. 23665533 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

With a reduction in maximum binding, an assay will report erroneously high values even in the absence of antigen and will exaggerate measurements of added antigen; with a reduction in affinity, the assay will under-report added antigen; in both cases there is a linear relationship between amount added and amount measured, with slope depending on the nature of the interference.

“With a reduction in maximum binding, an assay will report erroneously high values even in the absence of any antigen, and will exaggerate measurements of added antigen. The effect of a reduction in affinity is different; in this case, the assay will under‐report added antigen. In both cases, there will be a linear relationship between the amount of oxytocin added and the amount measured, although”

May combine two claims.

Bolus injections of oxytocin and vasopressin disappear rapidly, with an initial half-life that varies between species and with physiological state: in the rat both hormones disappear with an initial half-life of approximately 2 min, and in man oxytocin has a half-life of 4-10 min except in pregnancy when oxytocinase shortens it to 2-3 min.

“Bolus injections disappear rapidly, with an initial half‐life that varies between species and with physiological state: in the rat, both hormones disappear with an initial half‐life of approximately 2 min; in man, oxytocin has a half‐life of 4–10 min, except in pregnancy, when oxytocinase shortens this to 2–3 min.”

May combine two claims.

Later radioimmunoassay studies indicated that vasopressin disappeared from the circulation with a bi-exponential decay: the fast phase had a half-life of 5.4 min and a distribution volume close to the plasma volume, and the slower phase had a half-life of 78 min with a distribution volume close to the extracellular fluid volume.

“Later studies using radioimmunoassay indicated that vasopressin disappeared from the circulation with a bi‐exponential decay 107. The fast phase had a half‐life of 5.4 min and a distribution volume close to the plasma volume; the slower phase had a half‐life of 78 min with a distribution volume close to the extracellular fluid volume.”

• cites [107] Benmansour M, Rainfray M, Paillard F, Ardaillou R. Metabolic clearance rate of immunoreactive vasopressin in man. Eur J Clin Invest 1982; 12: 475– 480. 6818035 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Carter et al. claimed a biological validation for the ELISA in reporting that breastfeeding in women lowered salivary concentrations of oxytocin; because suckling-induced oxytocin release is indispensable for milk let-down in mammals, it is difficult to understand how such an anomalous observation qualifies as a validation.

“Finally, Carter et al. 119 claimed a biological validation for the ELISA in reporting that breastfeeding in women lowered salivary concentrations of oxytocin. Because suckling‐induced oxytocin release is indispensable for milk let‐down in mammals 123, it is difficult to understand how such an anomalous observation qualifies as a validation.”

• cites Carter et al. 119Not checked yet

  Not checked in the free run.

• cites [123] Russell JA, Leng G, Douglas AJ. The magnocellular oxytocin system, the fount of maternity: adaptations in pregnancy. Front Neuroendocrinol 2003; 24: 27– 61. 12609499 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

Plasma matrix proteins interact differently with different antibodies: there is no single radioimmunoassay and no single ELISA, and their susceptibility to plasma interference will vary with the antibody.

“Plasma matrix proteins interact differently with different antibodies: there is no single ‘radioimmunoassay’ and no single ‘ELISA’, and their susceptibility to plasma interference will vary with the antibody.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

In mice or rats, validation of vasopressin or oxytocin assays could be supported by measurements in gene knockout animals or hypophysectomised animals, where vasopressin and/or oxytocin are completely absent.

“In mice or rats, validation could be supported by measurements in gene knockout animals or hypophysectomised animals, where vasopressin and/or oxytocin are completely absent.”

May combine two claims.

Measuring oxytocin or vasopressin in cerebrospinal fluid, urine or saliva should have fewer problems because of the relative absence of proteins, but in urine high salt concentrations may cause problems.

“Measuring oxytocin or vasopressin in cerebrospinal fluid, urine or saliva should have fewer problems because of the relative absence of proteins but, in urine, high salt concentrations may cause problems.”

Classified as probably not a substantial claim. May combine two claims.

Plasma proteins differ from individual to individual and differ substantially from species to species, so any assay in unextracted plasma should be validated for the species in which it is measured and ideally in the individual in which it is measured.

“Plasma proteins differ from individual to individual and differ substantially from species to species, and so any assay in unextracted plasma should be validated for the species in which it is measured and, ideally, in the individual in which it is measured.”

• AssumptionRests on an assumption: Plasma proteins differ from individual to individual and differ substantially from species to species.

May combine two claims.

If the high levels arise from plasma protein interference, then these assays are, for unextracted plasma, no more than a random number generator, returning arbitrary values with a high variance and a skewed distribution.

“If so, these assays are, for unextracted plasma, no more than a random number generator, returning arbitrary values with a high variance and a skewed distribution.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Levels from an individual could be verified by constructing a standard curve in hormone-stripped plasma from that individual, although it would be impractical to do this for every individual sampled.

“Levels from an individual could be verified by constructing a standard curve in hormone‐stripped plasma from that individual 93, although it would be impractical to do this for every individual sampled.”

• cites [93] Henneberry HP, Slater JD, Eisen V, Fuhr S. Arginine vasopressin response to hypertonicity in hypertension studied by arginine vasopressin assay in unextracted plasma. J Hypertens 1992; 10: 221– 228. 1315818 sourceNot checked yet

  Not checked in the free run.

Classified as probably not a substantial claim.

The current impetus for measuring vasopressin and oxytocin in plasma derives from the assumption that such measurements are informative of the release of these peptides in the brain.

“The current impetus for measuring vasopressin and oxytocin in plasma derives from the assumption that such measurements are informative of the release of these peptides in the brain.”

Classified as probably not a substantial claim. May combine two claims.

There are no known conditions in which oxytocin is absent or grossly elevated in man; however, given the sensitivity of peripheral targets in man to very low concentrations, levels exceeding 10 pg/ml should be treated with scepticism.

“There are no known conditions in which oxytocin is absent or grossly elevated in man. However, given the sensitivity of peripheral targets in man to very low concentrations, levels exceeding 10 pg/ml should be treated with scepticism.”

• AssumptionRests on an assumption: There are no known conditions in which oxytocin is absent or grossly elevated in man.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Analytical verification by HPLC with chromatographic separation can confirm that measurements reflect the actual peptide, although care is needed to ensure that all high molecular weight proteins are eluted from the column and are checked for assay interference; there should be quantitative agreement between the amount of hormone in the appropriate fraction and the level reported by the assay.

“Analytical verification by HPLC with chromatographic separation 78 can confirm that measurements reflect the actual peptide, although care is needed to ensure that all high molecular weight proteins are eluted from the column and are checked for assay interference. There should be quantitative agreement between the amount of hormone in the appropriate fraction, and the level reported by the assay.”

• cites [78] Robertson GL, Klein LA, Roth J, Gorden P. Immunoassay of plasma vasopressin in man. Proc Natl Acad Sci USA 1970; 66: 1298– 1305. 5273455 PMC335819 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Interference will be apparent if, when samples are spiked with known amounts of hormone, the assay recovers amounts different to the expected amount; however, the absence of a difference is not conclusive because interference that reduces antibody affinity will lead to underestimation of added hormone, whereas interference that blocks antibody recognition will lead to overestimation, and both

“Such interference will be apparent if, when samples are spiked with known amounts of hormone, the assay recovers amounts different to the expected amount. However, the absence of a difference is not conclusive because interference that reduces antibody affinity will lead to underestimation of added hormone, whereas interference that blocks antibody recognition will lead to overestimation, and both”

May combine two claims.

Samples with high levels will not dilute in parallel to the standard curve if the levels arise from factors such as high salt levels or pH that interfere nonlinearly with antibody binding, although this test may not recognise protein interference that blocks antibody recognition of the antigen.

“Samples with high levels will not dilute in parallel to the standard curve if the levels arise from factors such as high salt levels or pH that interfere nonlinearly with antibody binding, although this test may not recognise protein interference that blocks antibody recognition of the antigen.”

Classified as probably not a substantial claim.

The fallacies in the supposition that plasma measurements are informative of brain peptide release have been discussed elsewhere.

“The fallacies in this supposition have been discussed elsewhere 2.”

• cites [2] Leng G, Ludwig M. Intranasal oxytocin: myths and delusions. Biol Psychiatry 2016; 79: 243– 250. 26049207 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

In a steady state, unbound oxytocin and vasopressin should distribute in extravascular fluids, and measurements in saliva might be useful for estimating steady-state secretion.

“In a steady‐state, unbound oxytocin and vasopressin should distribute in extravascular fluids, and measurements in saliva might be useful for estimating steady‐state secretion 119, 130, 131.”

• cites [119] Carter CS, Pournajafi‐Nazarloo H, Kramer KM, Ziegler TE, White‐Traut R, Bello D, Schwertz D. Oxytocin: behavioral associations and potential as a salivary biomarker. Ann N Y Acad Sci 2007; 1098: 312– 322. 17435137 sourceNot checked yet

  Not checked in the free run.

• cites [130] Jong TR, Menon R, Bludau A, Grund T, Biermeier V, Klampfl SM, Jurek B, Bosch OJ, Hellhammer J, Neumann ID. Salivary oxytocin concentrations in response to running, sexual self‐stimulation, breastfeeding and the TSST: the Regensburg Oxytocin Challenge (ROC) study. Psychoneuroendocrinology 2015; 62: 381– 388. 26385109 sourceNot checked yet

  Not checked in the free run.

• cites [131] Frijling JL, van Zuiden M, Nawijn L, Koch SB, Neumann ID, Veltman DJ, Olff M. Salivary oxytocin and vasopressin levels in police officers with and without post‐traumatic stress disorder. J Neuroendocrinol 2015; 27: 743– 751. 26184739 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Tsukazaki et al. developed an LC-MS/MS assay for vasopressin and showed that, correcting for plasma matrix effects, this assay gave close quantitative agreement with radioimmunoassay results from extracted human plasma samples, and had higher sensitivity and needed smaller sample volumes.

“Tsukazaki et al. 127 developed an LC‐MS/MS assay for vasopressin, and showed that, correcting for plasma matrix effects, this assay gave close quantitative agreement with radioimmunoassay results from extracted human plasma samples, and had higher sensitivity and needed smaller sample volumes.”

• cites Tsukazaki et al. 127Not checked yet

  Not checked in the free run.