Comparison between five methods for the separation of IgM.
Five serum fractionation methods were examined with regard to their suitability for use in the demonstration of specific IgM antibodies. It was found that Protein A - Sepharose CL - 4B chromatography leads to higher IgM recovery than absorption with whole Staphylococcus aureus (58% as opposed to 40%). It is also cheaper, since this absorbent can be reused. Contamination with 2-5% IgG and about 70% IgA is encountered with both methods. Quaternary aminoethyl (QAE) - Sephadex chromatography gives excellent IgM recovery (97%) and its cost and execution time are the lowest. IgG and IgA residues are 2% and 53% respectively. Ultrafiltration results in poorer recovery of IgM (31%). However, the sample volume can be brought back to its initial value, or concentrated still further. IgG and IgA residues are 2% and 14%. The cost of the method and the time needed for its execution are relatively high. Gel chromatography is the only method to give a completely IgG- and IgA-free IgM fraction. The percentage of IgM recovered varies in accordance with the ratio between the volume of the sample and that of the column. Very high values may be obtained, but at the expense of heavy dilution. The cost is low, but the method requires a considerable amount of time. In short, none of the five methods is clearly superior to the others. Whether or not a given method is chosen will thus depend on the relative importance attached to the percentage of IgM recovered, the presence or otherwise of IgA, sample dilution, cost, and rapidity of execution.
D002846:Chromatography, Affinity; D002847:Chromatography, Agarose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D007070:Immunoglobulin A / Q000032:analysis; D007074:Immunoglobulin G / Q000032:analysis; D007075:Immunoglobulin M / Q000302:isolation & purification*; D013205:Staphylococcal Protein A; D013211:Staphylococcus aureus / Q000276:immunology; D014462:Ultrafiltration
D007070:Immunoglobulin A; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D013205:Staphylococcal Protein A