A solid-phase protein assay: quantitation of protein in the nanogram range.
A solid-phase protein assay (SPA) for the determination of protein concentrations in the nanogram range is described. The technique is based on biotinylation of immobilized protein on the solid phase of a microtiter plate and quantitation of the protein-biotin complexes by peroxidase-coupled avidin. Compared to the routinely used Bradford and Lowry protein detection techniques, the described SPA assay is (depending upon the protein assayed) 1000 to 10,000 times more sensitive. Moreover, the SPA assay can be suitable for protein quantitation of samples containing agents which commonly interfere with the routinely used detection techniques. The SPA assay is a valuable addition to the Bradford and the Lowry techniques. Used in combination with an antigen-specific ELISA it gives a reliable ratio of specific versus total protein.
Sulter
M W
MW
Department of Dermatology, State University Hospital, Groningen, The Netherlands.
Kloosterhuis
G J
GJ
Coenraads
P J
PJ
Pas
H H
HH
D000818:Animals; D001360:Avidin; D001710:Biotin / Q000031:analogs & derivatives; D002417:Cattle; D002457:Cell Extracts / Q000032:analysis; D002623:Chemistry Techniques, Analytical / Q000379:methods; D003124:Colorimetry / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay; D005340:Fibrinogen / Q000032:analysis; D006735:Horseradish Peroxidase; D006801:Humans; D015603:Keratinocytes / Q000737:chemistry; D008832:Microchemistry / Q000379:methods; D010047:Ovalbumin / Q000032:analysis; D011506:Proteins / Q000032:analysis*; D012680:Sensitivity and Specificity; D012710:Serum Albumin, Bovine / Q000032:analysis; D013388:Succinimides
D002457:Cell Extracts; D011506:Proteins; D013388:Succinimides; C033788:biotinyl N-hydroxysuccinimide ester; D001360:Avidin; D012710:Serum Albumin, Bovine; D001710:Biotin; D005340:Fibrinogen; D010047:Ovalbumin; D006735:Horseradish Peroxidase