I am using native human ApoE derived from serum for cell culture studies. The company that I have ordered it from is not able to tell whether this protein is biologically active or not, because it has not been tested for functional studies. I have some data showing that there is some level of biological activity, however I am not sure if this is enough to continue my project. I was considering using recombinant ApoE, however the endotoxin level may skew my results, therefore I am trying to avoid switching to the recombinant ApoE. I would be grateful for any suggestions.
Ulrich is right, but a general test would be to show binding affinity to proteoglycans which can be done quickly with a heparin affinity column. You purchase columns from GE biosciences (or whoever they are now). Inactive will not bind well, but 'good' apoE should elute at about 250 mM NaCl.
Delipidated apoE (without any phospholipids or as part of a lipoprotein particle) will likely have different biological effects due to conformational differences of the protein when associated with other macromolecules. The test suggested by G. Shearer will at least inform you as to whether the heparin binding regions of apoE are exposed and functional and may provide some confidence that the protein is not irreversibly denatured
Send me an aliquot. I could do a cholesterol efflux and look whether or not it works as acceptor. Cholesterol efflux to apoE is mediated by ABCA1 at the cell membrane.
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