An immunoblotting procedure following agarose gel electrophoresis for detection of Bence Jones proteinuria compared with immunofixation and quantitative light chain determination.

Abstract

An immunoblotting procedure for the sensitive detection of Bence Jones proteinuria following agarose gel electrophoresis was developed. After immunonephelometric determination of urinary kappa and lambda light chains [employing antisera to human kappa and lambda light chains (free + bound)], urine samples (diluted to 2.5 mg/l kappa and lambda light chains, respectively) were electrophoretically separated using the Paragon system and blotted by capillary diffusion onto nitrocellulose. Rabbit anti-human kappa and lambda light chains reacted to kappa and lambda light chains attached to the membrane. Goat anti-rabbit IgG alkaline phosphatase conjugate was employed as detection system. The detection limit of the immunoblotting procedure (monoclonal component, as determined by serial dilutions) was 0.3 mg/l urine. Among 65 urine specimens received for routine testing for Bence Jones proteinuria, 32 monoclonal components (in 20 urine samples) were found by immunoblotting compared with 10 monoclonal components (in 9 urine samples) detected by immunofixation. In only 5 out of these 65 urine samples a kappa/lambda ratio (as determined immunonephelometrically) < 1 or > 5.2 (decision limits for discriminating between monoclonal and polyclonal urinary light chains; Boege F, Koehler B, Liebermann F. Eur J Clin Chem Clin Biochem 1990; 28:37-42) was observed. In conclusion, the immunoblotting method is superior to both immunofixation and immunonephelometry with respect to the diagnostic sensitivity for detection of Bence Jones proteinuria.