Hey, everyone!
Please help. I’ve been doing antibody analysis with one lambda beads for a few years. Several months ago we had noticed decreased MFI values of pos control. DTT treatment (ASHI procedure manual) significantly improved results with excellent correlation between FC/CDC XM data. The reason was presence of blocking anti-HLA and anti-idiotypic IgM. However, in almost 40% of my tests low bead counts were observed (<50) making the results unreportable to clinicians. Watching the procedure in details I had revealed formation of agglutinates after 1st washing of beads/serum mixture. This resulted in probe clogging and low bead counts consequently unless agglutinates were removed mechanically. IF anybody saw something like this and had an idea(s) how to circumvent this problem I will greatly appreciate,
Thank you very much.
Andrew

I NEVER use DTT for anything. It is very difficult to standardize and is more trouble than any good it can do. We have recently examined the effect of removal of IgM on IgG anti-HLA results with SA beads. We do not see any significant evidence of IgM blocking the IgG.

Furthermore, IgM anti-donor-HLA seem to be associated with loss of kidney and heart transplants.

Cheers,

-Peter Stastny