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Thromb Haemost 1985; 53(03): 377-380
DOI: 10.1055/s-0038-1661318
Original Article
Schattauer GmbH Stuttgart

Effect of Proteolysis on Quantitation of Plasma Fibronectin Concentration by Two Immunoassays (Electroimmunoassay and Immunoturbidimetric Technique)

K Bykowska
The Department of Radiobiology and Health Protection, Institute of Nuclear Chemistry and Technology, Warsaw, Poland
,
Z Wegrzynowicz
The Department of Radiobiology and Health Protection, Institute of Nuclear Chemistry and Technology, Warsaw, Poland
,
S Lopaciuk
The Department of Radiobiology and Health Protection, Institute of Nuclear Chemistry and Technology, Warsaw, Poland
,
M Kopeć
1   The Laboratory of Clinical Biochemistry, Institute of Haematology, Warsaw, Poland
› Author Affiliations
Further Information

Publication History

Received 19 September 1984

Accepted 20 March 1985

Publication Date:
21 August 2018 (online)

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Summary

Quantitation of fibronectin (FN) concentration is strongly influenced by FN fragmentation with trypsin, kallikrein and plasmin. Digestion by trypsin and kallikrein leads to a progressive decline in FN detectability by the immunoturbidimetric technique to zero values but is associated with an increase in the height of rockets in the Laurell’s electroimmunoassay. Plasmin mediated FN fragmentation induces a strong overestimation of the FN content by the electroimmunoassay and, at very high enzyme concentrations, provokes an underestimation of FN by the immunoturbidimetric technique. The decline in FN reactivity in the immunoturbidimetric assay coincides with the disappearance of heavy fractions migrating only slightly faster than native FN in SDS-PAGE. The increase in the height of rockets in the electroimmunoassay is the highest when fractions of intermediate rate of migration prevail in the SDS-PAGE pattern. Concomitant use of these two immunoassays can distinguish native FN from its degraded form and may possibly provide a partial explanation for discrepancies in published studies on the concentration of circulating FN in various pathological states.

Keywords

Fibronectin - Proteolysis - Immunoassays
 

  • References

  • 1 Saba TM, Jaffe E. Plasma fibronectin (opsonic glycoprotein): its synthesis by vascular endothelial cells and role in cardiopulmonary integrity after trauma as related to reticuloendothelial function. Am J Med 1980; 68: 577-594
    CrossrefPubMedSearch in Google Scholar
  • 2 Saba TM, Blumenstock FA, Weber P, Kaplan JE. Physiologic role for cold-insoluble globulin in systemic host defence. Implications of its characterization as the opsonic alfa-2-SB glycoprotein Ann NY Acad Sei 1978; 321: 43-55
    PubMedSearch in Google Scholar
  • 3 Blumenstock FA, Saba TM, Roccario E, Cho E, Kaplan JE. Opsonic fibronectin after trauma and particle injection determined by a peritoneal macrophage monolayer assay. J Reticuloendothel Soc 1981; 30: 61-71
    PubMedSearch in Google Scholar
  • 4 Mosher DF, Williams EM. Fibronectin concentration is decreased in plasma of severely ill patients with disseminated intravascular coagulation. J Lab Clin Med 1978; 91: 729-735
    PubMedSearch in Google Scholar
  • 5 Coulaud JM, Jacqueson A, Labrousse J, Allard D, Beyne P, Rapin J, Tenaillon A, Lissac J. Insuffisances respiratoires et circulatoires aiguës. Valeur pronostique du taux de fibronectine plasmatique Presse Med 1983; 12: 213-216
    PubMedSearch in Google Scholar
  • 6 Scovill WA, Saba TM, Kaplan JE. Deficit in reticuloendothelial humoral mechanisms after trauma. J Trauma 1976; 16: 898-904
    CrossrefPubMedSearch in Google Scholar
  • 7 Klingemann HG, Kosukavak M, Höfeier H, Havemann K. Fibronectin and factor VlII-related antigen in acute leukaemia. Hoppe-Seyler’s Z Physiol Chem 1983; 364: 269-277
    PubMedSearch in Google Scholar
  • 8 Reilly JT, Mackie MJ, McVerry BA. Observations on heterogeneity of fibronectin. Thromb Res 1984; 33: 289-295
    CrossrefPubMedSearch in Google Scholar
  • 9 Deno DC, McCafferty MH, Saba TM, Blumenstock FA. Mechanism of acute depletion of plasma fibronectin following thermal injury in rats. Appearance of a gelatin-like ligand in plasma J Clin Invest 1984; 73: 20-34
    CrossrefPubMedSearch in Google Scholar
  • 10 Lohmann J, Pott G. Qualitative and quantitative alterations of plasma fibronectin in patients suffering from shock septicemia. In: Factor XIII and fibronectin. Egbring R, Klingemann HG. (Eds) Die Medizinische Verlagsgesellschaft mbH; Marburg/Lahn: 1983. pp 257-261
    Search in Google Scholar
  • 11 Ruoslahti E, Hayman EG, Engvall E, Cothran WC, Butler WT. Alignment of biologically active domains in the fibronectin molecule. J Biol Chem 1981; 256: 7277-7281
    PubMedSearch in Google Scholar
  • 12 Amrani DL, Homandberg GA, Tooney NM, Wolfenstein-Todel C, Mosesson MW. Separation and analysis of the major forms of plasma fibronectin. Biochim Biophys Acta 1983; 748: 308-320
    CrossrefPubMedSearch in Google Scholar
  • 13 Miekka SI, Ingham KC, Menache D. Rapid methods for isolation of human plasma fibronectin. Thromb Res 1982; 27: 1-14
    CrossrefPubMedSearch in Google Scholar
  • 14 Mosesson MW, Umfleet RA. The cold insoluble globulin of human plasma. J Biol Chem 1970; 245: 5728-5736
    PubMedSearch in Google Scholar
  • 15 Laurell CB. Quantitative estimation of proteins by electrophoresis in agarose gel containing antibodies. Anal Biochem 1966; 15: 45-52
    CrossrefPubMedSearch in Google Scholar
  • 16 Saba TM, Albert WH, Blumenstock FA, Evanega G, Staehler F, Cho E. Evaluation of a rapid immunoturbidimetric assay for opsonic fibronectin in surgical and trauma patients. J Lab Clin Med 1981; 98: 482-491
    PubMedSearch in Google Scholar
  • 17 Swank RT, Munkers KD. Molecular weight analysis of oligopeptides by electrophoresis in polyacrylamide gel with sodium dodecyl sulfate. Anal Biochem 1971; 39: 462-477
    CrossrefPubMedSearch in Google Scholar
  • 18 Eriksen HO, Molke-Jensen F, Clemmensen I. Plasma fibronectin concentration in patients admitted to intensive care unit. Haematologia 1984; 17: 93-100
    PubMedSearch in Google Scholar
  • 19 Coulaud JM, Tenaillon A, Beyne P, Labrousse J, Lissac J, Rapin J, Jerome H, Salmona J-P, Jacqueson A, Allard D. Plasma fibronectin concentrations in critically ill patients. La Ricerca Clin Lab 1982; 12: 137-141
    PubMedSearch in Google Scholar
  • 20 Genestal M, Sié P, Hugot B, Cathala B, Boneu B, Lareng L. Fibronectine plasmatique. Rôle dans l’infection sévère en réanimation Presse Med 1983; 12: 1155-1158
    PubMedSearch in Google Scholar
  • 21 Choate JJ, Mosher DF. Fibronectin concentration in plasma of patients with breast cancer, colon cancer, and acute leukemia. Cancer 1983; 51: 1142-1147
    CrossrefPubMedSearch in Google Scholar
  • 22 Soria J, Soria C, Hourdille P, Sanchez R, Lafay P, Boisseau M. Fibronectin level in burned patients - confrontation with the level of other factors of coagulation - clinical application. In: Factor XIII and fibronectin. Egbring R, Klingemann HG. (Eds) Die Medizinische Verlagsgesellschaft; Marburg/Lahn: 1983. pp 245-254
    Search in Google Scholar
  • 23 Stathakis NE, Fountas A, Tsianos E. Plasma fibronectin in normal subjects and in various disease states. J Clin Pathol 1981; 34: 504-508
    CrossrefPubMedSearch in Google Scholar
  • 24 Grossman J, Pohlman T, Koerner F, Mosher D. Plasma fibronectin concentration in animal models of sepsis and endotoxemia. J Surg Res 1983; 34: 145-150
    CrossrefPubMedSearch in Google Scholar
  • 25 Schwarz HP, Aiginger P, Kuzmitz R. Fibronectin plasma concentration in patients with malignant diseases. In: Factor XIII and fibronectin. Egbring R, Klingemann HG. (Eds) Die Medizinische Verlagsgesellschaft; Marburg/Lahn: 1983. pp 263-267
    Search in Google Scholar
  • 26 Brühn HD, Heimburger N. Factor VIII-related antigen and cold insoluble globulin in leukemias and carcinomas. Haemostasis 1976; 5: 189-192
    PubMedSearch in Google Scholar
  • 27 Parsons RG, Todd HD, Kowal P. Isolation and identification of human serum fibronectin-like protein elevated during malignant disease. Cancer Res 1979; 39: 4341-4345
    PubMedSearch in Google Scholar
  • 28 Todd HD, Cofee MS, Waalkes TP, Abeloff MD, Parsons RG. Serum level of fibronectin and fibronectin-like DNA-binding protein in patients with various diseases. J Nat Cancer Inst 1980; 65: 901-904
    PubMedSearch in Google Scholar
  • 29 Gonzalez-Calvin J, Scully MF, Sanger Y, Fok J, Kakkar VV, Hughes RD, Gimson AE S, Williams R. Fibronectin in fulminant hepatic failure. B Med J 1982; 285: 1231-1232
    CrossrefPubMedSearch in Google Scholar
  • 30 Höfeler H, Klingemann HG, Egbring R. Fibronectin and factor VIIIR:Ag in acute and chronic hepatic failure. In: Factor XIII and fibronectin. Egbring R, Klingemann HG. (Eds) Die Medizinische Verlagsgesellschaft; Marburg/Lahn: 1983. pp 269-275
    Search in Google Scholar
  • 31 Gluud C, Dejgaard A, Clemmensen I. Plasma fibronectin concentrations in patients with liver diseases. Scand J Clin Lab Invest 1983; 43: 533-537
    CrossrefPubMedSearch in Google Scholar
  • 32 Matsuda M, Yamanaka T, Matsuda A. Distribution of fibronectin in plasma and liver in liver diseases. Clin Chim Acta 1982; 118: 191-199
    CrossrefPubMedSearch in Google Scholar
  • 33 Atichartakarn V, Marder VJ, Kirby EP, Budzynski AZ. Effects of enzymatic degradation on the subunit composition and biologic properties of human factor VIII. Blood 1978; 51: 281-291
    PubMedSearch in Google Scholar
  • 34 Kopeć M, Bykowska K, Lopaciuk S, Jelenska M, Kaczanowska J, Sopata I, Wojtecka E. Effects of neutral proteases from human leukocytes on structure and biological properties of human factor VIII. Thromb Haemostas 1980; 43: 211-217
    PubMedSearch in Google Scholar
  • 35 Selmer J, Eriksen H, Clemmensen I. Native and degraded fibronectin: new immunological methods for distinction. Scand J Clin Lab Invest 1983; 44: 57-63
    PubMedSearch in Google Scholar