From the crude polyclonal the crossreactive antibodies were extracted by incubation with Sepharose-bound human IgA and IgG.
Specific antibodies were absorpted by incubation with Sepharose-bound human IgM.
Specific antibodies were eluted by acidic buffer at pH 2.5 followed by neutralisation and dialysis.
After repeated binding with immobilized human IgM a minimum of 65% protein bound.
The purified polyclonal was conjugated to fluorescein isothiocyanate isomer I followed by gel-filtration and ion-exchange chromatography to clear unbound conjugate.
F/P ratio is 4-6 mol Fluorescein per 1 mol of goat IgM. There is low background fluorescence in normal tissue in contrast to cases of autoimmune desease where IgM is depostited.