Quantitative measurements of trefoil factor family peptides: Possibilities and pitfalls

Scandinavian Journal of Clinical and Laboratory Investigation

Volumn 73, Issue 3, 2013, Pages 193-202

  Samson, Mie Hessellund ^a  

^a AARHUS UNIVERSITY HOSPITAL   (Denmark)

Author keywords

ELISA; Immunoassay; TFF1; TFF2; TFF3

Indexed keywords

TREFOIL FACTOR FAMILY 1 PEPTIDE; TREFOIL FACTOR FAMILY 2 PEPTIDE; TREFOIL FACTOR FAMILY 3 PEPTIDE; TREFOIL PEPTIDE; UNCLASSIFIED DRUG;

EPITHELIUM; HUMAN; IMMUNOASSAY; INFLAMMATORY DISEASE; MEASUREMENT; MUCOSA; PREGNANCY; PRIORITY JOURNAL; PROTEIN SECRETION; QUANTITATIVE ANALYSIS; REVIEW; SURFACE PROPERTY;

AMINO ACID SEQUENCE; ENZYME-LINKED IMMUNOSORBENT ASSAY; EPITHELIAL CELLS; FEMALE; HUMANS; INFLAMMATION; MOLECULAR SEQUENCE DATA; MUCOUS MEMBRANE; NEOPLASMS; PEPTIDES; PREGNANCY; PROTEIN ISOFORMS;

LOTUS;

EID: 84876154817     PISSN: 00365513     EISSN: 15027686     Source Type: Journal    
DOI: 10.3109/00365513.2013.765962     Document Type: Review

References (91)

1. Kjellev S. The trefoil factor family-small peptides with multiple functionalities. Cell Mol Life Sci 2009; 66 : 1350-69
2. J ør gensen KH, T him L, J acobsen HE. P ancreatic spasmolytic polypeptide (PSP): I. Preparation and initial chemical characterization of a new polypeptide from porcine pancreas. Regul Pept 1982; 3 : 207-19
3. Jø r gensen K D, D iamant B, J ø r gensen K H, T him L. P ancreatic spasmolytic polypeptide (PSP): III. Pharmacology of a new porcine pancreatic polypeptide with spasmolytic and gastric acid secretion inhibitory effects. Regul Pept 1982; 3 : 231-43
4. Masiakowski P, B reathnach R, B loch J, G annon F, K rust A, Chambon P. Cloning of cDNA sequences of hormoneregulated genes from the MCF-7 human breast cancer cell line. Nucleic Acids Res 1982; 10 : 7895-903
5. Prud 'h omme JF, F ridlansky F, L e Cunff M, A tger M, Mercier-Bodart C, Pichon MF, Milgrom E. Cloning of a gene expressed in human breast cancer and regulated by estrogen in MCF-7 cells. DNA 1985; 4 : 11-21
6. Suemori S, L ynch-Devaney K, P odolsky DK. I dentifi cation and characterization of rat intestinal trefoil factor: Tissueand cell-specifi c member of the trefoil protein family. Proc Natl Acad Sci USA 1991; 88 : 11017-21
7. Tomasetto C, R io MC, G autier C, W olf C, H areuveni M, Chambon P, Lathe R. hSP, the domain-duplicated homolog of pS2 protein, is co-expressed with pS2 in stomach but not in breast carcinoma. EMBO J 1990; 9 : 407-14
8. Hauser F, P oulsom R, C hinery R, R ogers L A, H anby AM, Wright NA, Hoffmann W. hP1.B, a human P-domain peptide homologous with rat intestinal trefoil factor, is expressed also in the ulcer-Associated cell lineage and the uterus. Proc Natl Acad Sci USA 1993; 90 : 6961-5
9. Podolsky D K, L ynch-Devaney K, S tow J L, O ates P, M urgue B, deBeaumont M, Sands BE, Mahida YR. Identifi cation of human intestinal trefoil factor. Goblet cell-specifi c expression of a peptide targeted for apical secretion. J Biol Chem 1993; 268 : 6694-702
10. Thim L, May FEB. S tructure of mammalian trefoil factors and functional insights. Cell Mol Life Sci 2005; 62 : 2956-73
11. Thim L. A surprising sequence homology. B iochem J 1 988; 253 : 309
12. Hoffmann W. A new repetitive protein from X enopus laevis skin highly homologous to pancreatic spasmolytic polypeptide. J Biol Chem 1988; 263 : 7686-90
13. Baker ME. O estrogen-induced pS2 protein is similar to pancreatic spasmolytic polypeptide and the kringle domain. Biochem J 1988; 253 : 307-9
14. Wright NA, H offmann W, O tto WR, R io MC, T him L. R olling in the clover: Trefoil factor family (TFF)-domain peptides, cell migration and cancer. FEBS Lett 1997; 408 : 121-3
15. Gajhede M, P etersen TN, H enriksen A, P etersen J F, Dauter Z, Wilson KS, Thim L. Pancreatic spasmolytic polypeptide: Fi rst three-dimensional structure of a member of the mammalian trefoil family of peptides. Structure 1993; 1 : 253-62
16. Muskett FW, M ay FEB, W estley BR, F eeney J. S olution structure of the disulfi de-linked dimer of human intestinal trefoil factor (TFF3): The intermolecular orientation and interactions are markedly different from those of other dimeric trefoil proteins. Biochemistry 2003; 42 : 15139-47
17. Chinery R, Bates PA, De A, Freemont PS. C haracterisation of the single copy trefoil peptides intestinal trefoil factor and pS2 and their ability to form covalent dimers. FEBS Lett 1995; 357 : 50-4
18. Chadwick M P, W estley B R, M ay F E. H omodimerization and hetero-oligomerization of the single-domain trefoil protein pNR-2/pS2 through cysteine 58. Biochem J 1997; 327 : 117-23
19. Poulsen S S, T hulesen J, C hristensen L, N exo E, T him L. Metabolism of oral trefoil factor 2 (TFF2) and the effect of oral and parenteral TFF2 on gastric and duodenal ulcer healing in the rat. Gut 1999; 45 : 516-22
20. Kjellev S, V estergaard EM, N exo E, T hygesen P, E ghø j MS, Jeppesen PB, Thim L, Pedersen NB, Poulsen SS. Pharmacokinetics of trefoil peptides and their stability in gastrointestinal contents. Peptides 2007; 28 : 1197-206
21. Gouyer V, W iede A, B uisine MP, D ekeyser S, M oreau O, Lesuffl eur T, Hoffmann W, Huet G. Specifi c secretion of gel-forming mucins and TFF peptides in HT-29 cells of mucin-secreting phenotype. Biochim Biophys Acta 2001; 1539 : 71-84
22. Ota H, Hayama M, Momose M, El-Zimaity H MT, Matsuda K, Sano K, Maruta F, Okumura N, Katsuyama T. Colocalization of TFF2 with gland mucous cell mucin in gastric mucous cells and in extracellular mucous gel adherent to normal and damaged gastric mucosa. Histochem Cell Biol 2006; 126 : 617-25
23. Polshakov VI, W illiams MA, G argaro A R, F renkiel T A, Westley BR, Chadwick MP, May FE, Feeney J. High-resolution solution structure of human pNR-2/pS2: A single trefoil motif protein. J Mol Biol 1997; 267 : 418-32
24. Tomasetto C, M asson R, L inares J L, W endling C, L efebvre O, Chenard MP, Rio MC. pS2/TFF1 interacts directly with the VWFC cysteine-rich domains of mucins. Gastroenterology 2000; 118 : 70-80
25. Ruchaud-Sparagano MH, W estley BR, M ay FEB. The trefoil protein TFF1 is bound to MUC5AC in human gastric mucosa. Cell Mol Life Sci 2004; 61 : 1946-54
26. Kouznetsova I, Laubinger W, Kalbacher H, Kalinski T, Meyer F, Roessner A, Hoffmann W. Biosynthesis of gastrokine-2 in the human gastric mucosa: Restricted spatial expression along the antral gland axis and differential interaction with TFF1, TFF2 and mucins. Cell Physiol Biochem 2007; 20 : 899-908
27. Yu H, H e Y, Z hang X, P eng Z, Y ang Y, Z hu R, B ai J, T ian Y, Li X, Chen W, Fang D, Wang R. The rat IgGFc © BP and Muc2 C-Terminal domains and TFF3 in two intestinal mucus layers bind together by covalent interaction. PLoS ONE 2011; 6 : E20334
28. Thim L, M adsen F, P oulsen SS. E ffect of trefoil factors on the viscoelastic properties of mucus gels. Eur J Clin Invest 2002; 32 : 519-27
29. Kjellev S, N exo E, T him L, P oulsen SS. S ystemically administered trefoil factors are secreted into the gastric lumen and increase the viscosity of gastric contents. Br J Pharmacol 2006; 149 : 92-9
30. Dignass A, L ynch-Devaney K, K indon H, T him L, P odolsky DK. Trefoil peptides promote epithelial migration through a transforming growth factor beta-independent pathway. J Clin Invest 1994; 94 : 376-83
31. Playford RJ, M archbank T, C hinery R, E vison R, P ignatelli M, Boulton RA, Thim L, Hanby AM. Human spasmolytic polypeptide is a cytoprotective agent that stimulates cell migration. Gastroenterology 1995; 108 : 108-16
32. Oertel M, G raness A, T him L, B ü hling F, K albacher H, Hoffmann W. Trefoil factor family-peptides promote migration of human bronchial epithelial cells: Synergistic effect with epidermal growth factor. Am J Respir Cell Mol Biol 2001; 25 : 418-24
33. May F EB, S emple J I, P rest S J, W estley B R. E xpression and motogenic activity of TFF2 in human breast cancer cells. Peptides 2004; 25 : 865-72
34. Storesund T, H ayashi K, K olltveit K M, B ryne M, S chenck K. Salivary trefoil factor 3 enhances migration of oral keratinocytes. Eur J Oral Sci 2008; 116 : 135-40
35. Lalani EN, W illiams R, J ayaram Y, G ilbert C, C haudhary KS, Siu LS, Koumarianou A, Playford R, Stamp GW. Trefoil factor-2, human spasmolytic polypeptide, promotes branching morphogenesis in MCF-7 cells. Lab Invest 1999; 79 : 537-46
36. Taupin DR, K inoshita K, P odolsky DK. I ntestinal trefoil factor confers colonic epithelial resistance to apoptosis. Proc Natl Acad Sci USA 2000; 97 : 799-804
37. Siu LS, R omanska H, A bel PD, B aus-Loncar M, K ayademir T, Stamp GWH, Lalani EN. TFF2 (trefoil family factor2) inhibits apoptosis in breast and colorectal cancer cell lines. Peptides 2004; 25 : 855-63
38. Buron N, G uery L, C reuzot-Garcher C, L afontaine P O, Bron A, Rio MC, Solary E. Trefoil factor TFF1-induced protection of conjunctival cells from apoptosis at premitochondrial and postmitochondrial levels. Invest Ophthalmol Vis Sci 2008; 49 : 3790-8
39. Rodrigues S, V an Aken E, V an Bocxlaer S, A ttoub S, N guyen QD, Bruyneel E, Westley BR, May FEB, Thim L, Mareel M, Gespach C, Emami S. Trefoil peptides as proangiogenic factors in vivo and in vitro: Implication of cyclooxygenase-2 and EGF receptor signaling. FASEB J 2003; 17 : 7-16
40. Baus-Loncar M, K ayademir T, T akaishi S, W ang T. T refoil factor family 2 defi ciency and immune response. Cell Mol Life Sci 2005; 62 : 2947-55
41. Dubeykovskaya Z, D ubeykovskiy A, S olal-Cohen J, W ang TC. Secreted trefoil factor 2 activates the CXCR4 receptor in epithelial and lymphocytic cancer cell lines. J Biol Chem 2009; 284 : 3650-62
42. Probst J C, Z etzsche T, W eber M, T heilemann P, S kutella T, Landgraf R, Jirikowski GF. Human intestinal trefoil factor is expressed in human hypothalamus and pituitary: Evidence for a novel neuropeptide. FASEB J 1996; 10 : 1518-23
43. Jackerott M, L ee YC, M øl lg år d K, K ofod H, J ensen J, Rohleder S, Neubauer N, Gaarn LW, Lykke J, Dodge R, Dalgaard LT, S ø strup B, Jensen DB, Thim L, Nexo E, Thams P, Bisgaard HC, Nielsen JH. Trefoil factors are expressed in human and rat endocrine pancreas: Differential regulation by growth hormone. Endocrinology 2006; 147 : 5752-9
44. Mahmood A, M elley L, F itzGerald A J, G hosh S, P layford RJ. Trial of trefoil factor 3 enemas, in combination with oral 5-Aminosalicylic acid, for the treatment of mild-To-moderate left-sided ulcerative colitis. Aliment Pharmacol Ther 2005; 21 : 1357-64
45. Peterson D E, B arker N P, A khmadullina L I, R odionova I, Sherman NZ, Davidenko IS, Rakovskaya GN, Gotovkin EA, Shinkarev SA, Kopp MV, Kulikov EP, Moiseyenko VM, Gertner JM, Firsov I, Tuleneva T, Yarosh A, Woon CW. Phase II, randomized, double-blind, placebo-controlled study of recombinant human intestinal trefoil factor oral spray for prevention of oral mucositis in patients with colorectal cancer who are receiving fl uorouracil-based chemotherapy. J Clin Oncol 2009; 27 : 4333-8
46. May FEB. The potential of trefoil proteins as biomarkers in human cancer. Biomark Med 2012; 6 : 301-4
47. Foekens J A, R io M C, S eguin P, v an Putten W L, F auque J, Nap M, Klijn JG, Chambon P. Prediction of relapse and survival in breast cancer patients by pS2 protein status. Cancer Res 1990; 50 : 3832-7
48. Predine J, S pyratos F, P rud 'h omme JF, A ndrieu C, H acene K, Brunet M, Pallud C, Milgrom E. Enzyme-linked immunosorbent assay of pS2 in breast cancers, benign tumors, and normal breast tissues. Correlation with prognosis and adjuvant hormone therapy. Cancer 1992; 69 : 2116-23
49. Miyashita S, N omoto H, K onishi H, H ayashi K. E stimation of pS2 protein level in human body fl uids by a sensitive twosite enzyme immunoassay. Clin Chim Acta 1994; 228 : 71-81
50. Z ava DT, D uwe G. E strogenic and antiproliferative properties of genistein and other fl avonoids in human breast cancer cells in vitro. Nutr Cancer 1997; 27 : 31-40
51. Rosenberg Zand RS, J enkins DJ, D iamandis EP. D evelopment and evaluation of a competitive time-resolved immunofl uorometric assay for the estrogen-regulated protein pS2. J Clin Lab Anal 1999; 13 : 241-5
52. Taupin D, Pedersen J, F amilari M, C ook G, Y eomans N, Giraud AS. Augmented intestinal trefoil factor (TFF3) and loss of pS2 (TFF1) expression precedes metaplastic differentiation of gastric epithelium. Lab Invest 2001; 81 : 397-408
53. Vestergaard E M, Brynskov J, Ejskjaer K, Clausen JT, T him L, Nexo E, Poulsen SS. Immunoassays of human trefoil factors 1 and 2: Measured on serum from patients with infl ammatory bowel disease. Scand J Clin Lab Invest 2004; 64 : 146-56
54. Pu M, M ay FEB, P layford RJ, R uchaud-Sparagano MH, Westley BR. Development of a two-site ELISA assay for the dimeric form of human TFF1. Peptides 2004; 25 : 731-6
55. Kaise M, M iwa J, T ashiro J, O hmoto Y, M orimoto S, K ato M, Urashima M, Ikegami M, Tajiri H. The combination of serum trefoil factor 3 and pepsinogen testing is a valid nonendoscopic biomarker for predicting the presence of gastric cancer: A new marker for gastric cancer risk. J Gastroenterol 2011; 46 : 736-45
56. Qu Y, Yang Y, Ma D, Xiao W. I ncreased trefoil factor 3 levels in the serum of patients with three major histological subtypes of lung cancer. Oncol Rep 2012; 27 : 1277-83
57. Babyatsky MW, d eBeaumont M, T him L, P odolsky DK. Oral trefoil peptides protect against ethanol-And indomethacin-induced gastric injury in rats. Gastroenterology 1996; 110 : 489-97
58. Tu SP, C hi AL, A i W, T akaishi S, D ubeykovskaya Z, Q uante M, Fox JG, Wang TC. p53 inhibition of AP1-dependent TFF2 expression induces apoptosis and inhibits cell migration in gastric cancer cells. Am J Physiol Gastrointest Liver Physiol 2009; 297 : G385-96
59. Kubota S, Y amauchi K, S ugano M, K awasaki K, S ugiyama A, Matsuzawa K, Akamatsu T, Ohmoto Y, Ota H. Pathophysiological investigation of the gastric surface mucous gel layer of patients with Helicobacter pylori infection by using immunoassays for trefoil factor family 2 and gastric gland mucous celltype mucin in gastric juice. Dig Dis Sci 2011; 56 : 3498-506
60. Taupin DR, P ang KC, G reen SP, Giraud A S. The trefoil peptides spasmolytic polypeptide and intestinal trefoil factor are major secretory products of the rat gut. Peptides 1995; 16 : 1001-5
61. Moro F, L evenez F, Durual S, Plaisancié P, T him L, Giraud AS, Cuber JC. Secretion of the trefoil factor TFF3 from the isolated vascularly perfused rat colon. Regul Pept; 101 : 35-41
62. Vestergaard EM, P oulsen SS, G r øn baek H, L arsen R, Nielsen AM, Ejskjaer K, Clausen JT, Thim L, Nexo E. Development and evaluation of an ELISA for human trefoil factor 3. Clin Chem 2002; 48 : 1689-95
63. Bignotti E, R avaggi A, T assi R A, C alza S, R ossi E, F alchetti M, Romani C, Bandiera E, Odicino FE, Pecorelli S, Santin AD. Trefoil factor 3: A novel serum marker identifi ed by gene expression profi ling in high-grade endometrial carcinomas. Br J Cancer 2008; 99 : 768-73
64. Xue H, L ü B, Z hang J, W u M, H uang Q, W u Q, S heng H, Wu D, Hu J, Lai M. Identifi cation of serum biomarkers for colorectal cancer metastasis using a differential secretome approach. J Proteome Res 2010; 9 : 545-55
65. Astor B C, K ö t tgen A, H wang S-J, B havsar N, F ox CS, Coresh J. Trefoil factor 3 predicts incident chronic kidney disease: A case-control study nested within the Atherosclerosis Risk in Communities (ARIC) Study. Am J Nephrol 2011; 34 : 291-7
66. Schulze U, H ampel U, S el S, G oecke TW, T h äl e V, G arreis F, Paulsen F. Fresh and cryopreserved amniotic membrane secrete the trefoil factor family peptide 3 that is well known to promote wound healing. Histochem Cell Biol 2012; 138 : 243-50
67. Samson M H, N exo E. V alidation of commercial assays for measurements of trefoil factor family peptides in serum. Clin Chem Lab Med 2011; 49 : 2057-60
68. Newton JL, A llen A, W estley BR, M ay FE. The human trefoil peptide, TFF1, is present in different molecular forms that are intimately associated with mucus in normal stomach. Gut 2000; 46 : 312-20
69. Westley BR, G riffi n SM, M ay FEB. I nteraction between TFF1, a gastric tumor suppressor trefoil protein, and TFIZ1, a brichos domain-containing protein with homology to SP-C. Biochemistry 2005; 44 : 7967-75
70. Tosco A, M onti M C, F ontanella B, R io M-C, G omez-Paloma L, Leone A, Marzullo L. Copper-binding activity of Trefoil factor 1 (TFF1): A new perspective in the study of the multifunctional roles of TFFs. Peptides 2007; 28 : 1461-9
71. Tosco A, M onti MC, F ontanella B, M ontefusco S, D 'A ndrea L, Ziaco B, Baldantoni D, Rio MC, Marzullo L. Copper binds the carboxy-Terminus of trefoil protein 1 (TFF1), favoring its homodimerization and motogenic activity. Cell Mol Life Sci 2010; 67 : 1943-55
72. May F E, S emple J I, N ewton J L, W estley B R. The human two domain trefoil protein, TFF2, is glycosylated in vivo in the stomach. Gut 2000; 46 : 454-9
73. Kouznetsova I, P eitz U, V ieth M, M eyer F, V estergaard EM, Malfertheiner P, Roessner A, Lippert H, Hoffmann W. A gradient of TFF3 (trefoil factor family 3) peptide synthesis within the normal human gastric mucosa. Cell Tissue Res 2004; 316 : 155-65
74. Albert TK, L aubinger W, M ül ler S, H anisch F-G, K alinski T, Meyer F, Hoffmann W. Human intestinal TFF3 forms disulfi de-linked heteromers with the mucus-Associated FCGBP protein and is released by hydrogen sulfi de. J Proteome Res 2010; 9 : 3108-17
75. Samson MH, Chaiyarit P, Nortvig H, Vestergaard EM, Ernst E, Nexo E. Trefoil factor family peptides in human saliva and cyclical cervical mucus. Method evaluation and results on healthy individuals. Clin Chem Lab Med 2011; 49 : 861-8
76. Samson MH, Vestergaard EM, Milman N, Poulsen SS, Nexo E. Circulating serum trefoil factors increase dramatically during pregnancy. Scand J Clin Lab Invest 2008; 68 : 369-74
77. Samson M H, P oulsen S S, O beid R, H errmann W, N exo E. Trefoil factor family peptides in the human foetus and at birth. Eur J Clin Invest 2011; 41 : 785-92
78. Vestergaard E M, N exo E, W endt A, G uthmann F. T refoil factors in human milk. Early Hum Dev 2008; 84 : 631-5
79. Madsen J, N ielsen O, T orn øe I, T him L, H olmskov U. T issue localization of human trefoil factors 1, 2, and 3. J Histochem Cytochem 2007; 55 : 505-13
80. Chenard M P, T omasetto C, B ellocq J P, R io M C. U rinary pS2/TFF1 levels in the management of hormonodependent breast carcinomas. Peptides 2004; 25 : 737-43
81. Higashiyama M, D oi O, K odama K, Y okuchi H, I naji H, Tateishi R. Estimation of serum level of pS2 protein in patients with lung adenocarcinoma. Anticancer Res 1996 ; 16 : 2351-5
82. Aikou S, Ohmoto Y, G unji T, M atsuhashi N, O htsu H, M iura H, Kubota K, Yamagata Y, Seto Y, Nakajima A, Goldenring JR, Kaminishi M, Nomura S. Tests for serum levels of trefoil factor family proteins can improve gastric cancer screening. Gastroenterology 2011 ; 141 : 837-845. e1-7
83. Kaise M, M iwa J, F ujimoto A, T ashiro J, T agami D, S ano H, Ohmoto Y. Influence of Helicobacter pylori status and eradication on the serum levels of trefoil factors and pepsinogen test: Serum trefoil factor 3 is a stable biomarker. Gastric Cancer 2012Aug. 21
84. Vestergaard E M, B orre M, P oulsen S S, N exo E, T ør ring N. Plasma levels of trefoil factors are increased in patients with advanced prostate cancer. Clin Cancer Res 2006; 12 : 807-12
85. Vestergaard E M, N exo E, T ør ring N, B orre M, Ø r ntoft TF, S ø rensen KD. Promoter hypomethylation and upregulation of trefoil factors in prostate cancer. Int J Cancer 2010; 127 : 1857-65
86. Chaiyarit P, U trawichian A, L eelayuwat C, V atanasapt P, Chanchareonsook N, Samson MH, Giraud AS. Investigation of trefoil factor expression in saliva and oral mucosal tissues of patients with oral squamous cell carcinoma. Clin Oral Investig 2012; 16 : 1549-56
87. Babyatsky M, Lin J, Yio X, Chen A, Zhang JY, Z heng Y, Twyman C, Bao X, Schwartz M, Thung S, Lawrence Werther J, Itzkowitz S. Trefoil factor-3 expression in human colon cancer liver metastasis. Clin Exp Metastasis 2009; 26 : 143-51
88. Gr øn baek H, V estergaard EM, H ey H, N ielsen JN, N exo E. Serum trefoil factors in patients with infl ammatory bowel disease. Digestion 2006; 74 : 33-9
89. Chaiyarit P, C hayasadom A, W ara-Aswapati N, H ormdee D, Sittisomwong S, Nakaresisoon S, Samson MH, Pitiphat W, Giraud AS. Trefoil factors in saliva and gingival tissues of patients with chronic periodontitis. J Periodontol 2012; 83 : 1129-38
90. Verey F, N exo E, G reenwood R, B erry M, C orfi eld AP. T refoil factor family peptides are increased in the saliva of children with mucositis. Clin Chem Lab Med 2011; 49 : 2051-5
91. Goussard J, L echevrel C, R oussel G, C ren H, B era O, S ala M. Immunoradiometric assay of pS2 protein in breast cancer cytosols. Clin Chem 1991; 37 : 1759-62