Recombinant human heart-type fatty acid-binding protein as standard in immunochemical assays.
Cytoplasmic heart-type fatty acid-binding protein has recently gained much attention in clinical diagnosis as a very early marker of acute myocardial infarction. Immunoassays have been developed for determination of this protein in plasma and urine samples. In the present study it is shown that those types of fatty acid-binding proteins which are abundant in tissues other than heart and muscle do not interfere with immunochemical determination of heart-type fatty acid-binding protein. To provide sufficient protein of consistent quality as standard in these immunoassays, human heart-type fatty acid-binding protein was cloned, expressed in Escherichia coli and purified to homogeneity. For quantitation of the recombinant protein its extinction coefficient was determined. Comparison of the recombinant and tissue-derived proteins by a variety of methods revealed both proteins to show similar kinetic as well as equilibrium constants with respect to two monoclonal antibodies currently applied in immunochemical detection of heart-type fatty acid-binding protein. Both preparations were indistiguishable in sandwich-ELISA and immunosensor measurements. A high stability of the recombinant protein was proven by ELISA measurements during storage and several freeze and thaw cycles. Thus, recombinant and tissue-derived heart-type fatty acid-binding proteins are immunochemically equivalent. The recombinant human heart-type fatty acid-binding protein is now available as standard for immunoassays.
Institut für Chemo- und Biosensorik, Münster, Germany.
A
A
Schreiber
B
B
Specht
M M
MM
Pelsers
J F
JF
Glatz
T
T
Börchers
F
F
Spener
D000911:Antibodies, Monoclonal; D000937:Antigen-Antibody Reactions; D002352:Carrier Proteins / Q000032:analysis* / Q000235:genetics / Q000592:standards*; D003001:Cloning, Molecular; D003429:Cross Reactions; D004355:Drug Stability; D004797:Enzyme-Linked Immunosorbent Assay / Q000592:standards; D004926:Escherichia coli / Q000235:genetics; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids / Q000378:metabolism*; D006801:Humans; D007120:Immunochemistry / Q000592:standards*; D066298:In Vitro Techniques; D007700:Kinetics; D018996:Myelin P2 Protein / Q000032:analysis* / Q000235:genetics / Q000592:standards*; D009203:Myocardial Infarction / Q000175:diagnosis / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism*; D009363:Neoplasm Proteins; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics / Q000592:standards; D012015:Reference Standards; D025521:Tumor Suppressor Proteins
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; C107608:FABP7 protein, human; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D005227:Fatty Acids; D018996:Myelin P2 Protein; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D025521:Tumor Suppressor Proteins