While performing the enzyme activity of XR and XDH at 340 nm, during time scan of 3 mins, my readings fluctuate invariably. Is there any certain factor to be taken care of while putting up the enzyme reaction. I am following this paper:
S.I. Yokoyama, T. Suzuki, K. Kawai, H. Horitsu, K. Takamizawa (1995) Purification, Characterization and Structure Analysis of NADPH-Dependent D-Xylose Reductase from Candida tropicalis. J. Ferm. Bioeng. 79 (3) 217-223.
Dear Shalley,
I would suggest the use of a cell stirring magnetic device, so the contents of the cell (cuvette) are constantly mixed and therefore you are constantly measuring a more homogeneous solution.
Kind regards,
Ade Kujore
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Additionally, variation can result from air bubbles forming in the solution during the reaction. This is most likely to occur if the reaction is performed at an elevated temperature. Pre-warming the solutions before starting the reaction will help. Degassing, if it is feasible, will also help.
Watch out for particles of dust and debris in your solutions. Filter stock solutions, if necessary, to remove it.
Thanks Ade and Adam. I tried both the suggestions but with XR I am still facing the same problem as readings are going down in the negative range. But with XDH, results are consistent.
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