Sample preparation is an important element in the detection of toxins in food samples. In this work, a simple analytical sample preparation method for recovery of small amount of staphylococcal enterotoxin B (SEB) and staphylococcal enterotoxin A (SEA) in food samples was developed. Cation exchanger carboxymethylcellulose (CM) was used for small-scale batch chromatography isolation of SEB from infant formula and from mushrooms spiked with SEB. The resulting materials were analyzed for SEB by Western immunoblotting. Nearly all of the extraneous substances in the sample were removed by this procedure with no significant loss of the toxin. Using this method, even small amounts of SE (0.75 ng/g) can be recovered and immunologically analyzed by Western blotting or by ELISA with a very low background. Because this method is effective, rapid, simple and inexpensive, it has the potential to be a general method for the preparation of samples used for analysis of SEs.