These tools will no longer be maintained as of December 31, 2024. Archived website can be found here. PubMed4Hh GitHub repository can be found here. Contact NLM Customer Service if you have questions.


Pubmed for Handhelds

PUBMED FOR HANDHELDS

Search MEDLINE/PubMed

  • Title: Measurement of carbonic anhydrase activity using a sensitive fluorometric assay.
    Author: Shingles R, Moroney JV.
    Journal: Anal Biochem; 1997 Oct 01; 252(1):190-7. PubMed ID: 9324959.
    Abstract:
    The dehydration reaction of bicarbonate was measured using the fluorescent pH indicator, 8-hydroxypyrene-1,3,6-trisulfonate (pyranine), in combination with stopped-flow spectrofluorometry. The initial rate of bicarbonate dehydration was measured after mixing a pH 6.0 solution with a pH 8.0 solution containing bicarbonate. Addition of carbonic anhydrase to the pH 6.0 solution enabled the measurement of the initial rate of activity at physiological temperatures with resolution times of 2 ms. This assay was used to resolve differences in activity and sensitivity to sulfonamides by comparing mammalian carbonic anhydrase isoforms. The fluorescent technique used in this study is very sensitive, allowing the determination of initial rates with a protein concentration as little as 65 ng/ml. Pyranine can also be loaded into membrane vesicles to follow carbonic anhydrase activity within vesicles. The change in pH within vesicles is dependent on the concentration of externally added bicarbonate and the presence of carbonic anhydrase on either side of the membrane. Therefore, this assay can be used to measure carbon dioxide flux across membranes and to assess the contribution of carbonic anhydrase to this flux.
    [Abstract] [Full Text] [Related] [New Search]