Precolumn affinity capillary electrophoresis for the identification of clinically relevant proteins in human serum: application to human cardiac troponin I.
An approach has been developed to the on-line extraction and identification of clinical disease-state marker proteins in human serum. Fabrication of capillaries with integral packed beds for the online determination of human cardiac troponin I (cTnI), a diagnostic marker for myocardial infarction, at clinically relevant levels (2 nmol/L) in serum is demonstrated. The technique, termed precolumn affinity capillary electrophoresis (PA-CE), utilizes a short (approximately 5 mm) packed bed of porous silica containing covalently immobilized monoclonal anti-cTnI antibodies directly integrated within a separation capillary for the selective retention of cTnI from a complex matrix. Following a rinsing step to eliminate nonspecifically bound serum proteins and other impurities from the column, desorption of the antigen into the separation region of the PA-CE capillary for subsequent measurement of femto-molar amounts of cTnI by CE is effected by the injection of an appropriate elution buffer. Advantages of this approach over previously reported affinity preconcentration techniques, related applications for PA-CE technology, and its potential for use in the development of a certified reference material for cTnI in serum are discussed.
Dalluge
J J
JJ
Analytical Chemistry Division, National Institute of Standards and Technology, Gaithersburg, Maryland 20899-0001, USA. joseph.dalluge@nist.gov
Sander
L C
LC
D001798:Blood Proteins / Q000032:analysis*; D019075:Electrophoresis, Capillary; D006801:Humans; D009206:Myocardium / Q000737:chemistry*; D019210:Troponin I / Q000032:analysis*
D001798:Blood Proteins; D019210:Troponin I