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I'm doing a project on the purification of sheep platelet cyclooxygenase and have read a paper. But there are some steps of the protocol which I don't understand. First, hydrophobic chromatography using ibuprofen-sepharose affinity column was used. Next, fractions were subjected to 2 metal-chelate chromatographies - IDA/Zn column then TED/Zn column. Cyclooxygenase binded to IDA/Zn which was eluted with imidazole. The cyclooxygenase-containing fractions were put through the TED/Zn column but cyclooxygenase did not bind and was collected in the unbound effluent. The unbound effluent was then put through a Haemin-Sepharose affinity chromatography column and purified cyclooxygenase was obtained. Can anyone pls enlighten me about why a series of affinity chromatography was used and what proteins were gotten rid of in each step? IDA= iminodiacetic acid; TED= Tris(carboxymethyl)ethylenediamine. Thanks.