I am currently using a pair of matched anti mouse IL-5 antibodies (TRFK
4 and TRFK 5 monoclonals) in a sandwich style ELISA to detect IL-5 in
the cell supernatants of MC/9 cells. I am concerned that I may be
measuring only a small percentage of the total IL-5 present in the
samples due to interference from soluble IL-5 receptors and/or serum
proteins (ie: the epitopes on IL-5 recognized by either the capture or
detection antibody may be masked).
I have encountered this problem in the past when measuring TNF, but was
able to solve it by using a polyclonal based competitive EIA. To my
surprise the typical sandwich ELISA measured only 10 % of the TNF
detected by the competitive EIA.
Unfortunately, I have been unable to source a polyclonal based
competitive EIA specific for mouse IL-5 and am hesitant to purchase the
antibody and produce my own. Does anyone have any recommendations on how
to quantitatively measure total IL-5 in cell supernatant without
interference from soluble receptors or other proteins that may bind?
