By Marlene A. DeLuca (Eds.)
The severely acclaimed laboratory general, Methods in Enzymology, is among the such a lot hugely revered guides within the box of biochemistry. in view that 1955, each one quantity has been eagerly awaited, often consulted, and praised through researchers and reviewers alike. The sequence includes a lot fabric nonetheless appropriate at the present time - really a necessary book for researchers in all fields of lifestyles sciences
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Extra info for Bioluminescence and Chemiluminescence
3] PREPARATION OF PARTIALLY PURIFIED FIREFLY LUCIFERASE 31 mortar with a little acid-washed sand. Extraction buffer (20 ml per gram of tails) is added gradually with continued grinding. 7. The activity of the firefly tails and the efficiency of the grinding may be judged from the light emitted by the suspension. , 15,000 rpm for 30 min, and the clear supernatant is isolated with minimal contamination from the upper lipid layer. One gram of firefly tails is usually extracted in a single operation.
4. The hydrogen chloride is first dried by passage through concentrated H2SO4. The safety bottle trap is included as a precaution against "sucking back" of contents of the reaction vessel (l-liter three-necked flask). Hydrogen chloride generated in this manner is passed over the surface drying tube nc. H2SO 4 ~ - ' - / ' ~ " NH 4 Cl ~\conc. - 7~ eter safety H2SO4 battle trap heating mantle FIG. 4. Apparatus for the in situ production of pyridine hydrochloride and demethylation of 2-cyano-6-methoxybenzothiazole.
The ion-exchange chromatographic method of Btny and Dolivo 3 requires 3 days' chromatography. In the publication, the purification is evaluated in a manner different from the present one, but it is probably of the same order of magnitude. The stepwise elution procedure mentioned in the section Comments on the Purification Procedure gives the luciferase in a very small volume. It may be advantageous if a less efficient purification, in particular with respect to nucleosidediphosphate kinase, can be tolerated.
Bioluminescence and Chemiluminescence by Marlene A. DeLuca (Eds.)