Plasma interference in an enzyme-linked immunosorbant assay using a commercial matched antibody pair.
In this study, severe plasma interference was repeatedly documented in an IL-1ra sandwich enzyme-linked immunosorbant assay (ELISA) using a commercial matched antibody pair. Several physical and biochemical treatments were used in an attempt to alleviate this plasma effect including the following: buffer optimization, sample dilution, increasing incubation temperature, heat treatment of plasma, increasing detergent concentrations, glutaraldehyde pretreatment of the plate and the addition of polyethylene glycol (PEG). Evaluation of several buffers demonstrated that the range of optical densities could be increased dramatically with the use of an appropriate buffer. Of the treatments examined, only the addition of polyethylene glycol (PEG) to the dilution buffer created a marked improvement in the ELISA, despite a resulting background increase. Further investigation demonstrated that 10% PEG in the dilution buffer added to biotinylated antibody and the streptavidin provided the greatest improvement to the sensitivity of the ELISA.
Nemzek
J A
JA
The University of Michigan, Department of Pathology, Ann Arbor, Michigan 48109-0602, USA.
Newcomb
D E
DE
Call
D R
DR
Remick
D G
DG
D000906:Antibodies / Q000276:immunology; D002021:Buffers; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D053590:Interleukin 1 Receptor Antagonist Protein; D010949:Plasma / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000494:pharmacology; D011786:Quality Control; D011994:Recombinant Proteins / Q000032:analysis; D012795:Sialoglycoproteins / Q000032:analysis*
D000906:Antibodies; D002021:Buffers; C507763:IL1RN protein, human; D053590:Interleukin 1 Receptor Antagonist Protein; D011994:Recombinant Proteins; D012795:Sialoglycoproteins; D011092:Polyethylene Glycols