An in situ study of beta-glucosidase activity in normal and Gaucher fibroblasts with fluorogenic probes.
AuthorsKohen E, Kohen C, Hirschberg JG, Santus R, Grabowski G, Mangel W, Gatt S, Prince J
JournalCell Biochem Funct
PubMed ID8403230
'Beta-glucosidase activity was evaluated in situ by means of fluorogenic probes in normal human fibroblasts and fibroblasts from homozygous carriers of the Gaucher trait. Probe internalization, targeting to lysosomes and post-cleavage probe retention were the primary concerns. Internalization and targeting were attempted by in situ photosensitized labilization of lysosomal membranes, ... More
Yeast exo-beta-glucanases can be used as efficient and readily detectable reporter genes in Saccharomyces cerevisiae.
AuthorsCid VJ, Alvarez AM, Santos AI, Nombela C, Sanchez M
JournalYeast
PubMed ID7975893
'Yeast exo-1,3-beta-glucanases are secretable proteins whose function is basically trophic and may also be involved in cell wall glucan hydrolytic processes. Since fluorescein di(beta-D-glucopyranoside) is a fluorogenic substrate detectable and quantifiable by flow cytometry, it was used for testing the ability of the EXG1 gene product of Saccharomyces cerevisiae and ... More
A flow cytometric assay for lysosomal glucocerebrosidase.
Authorsvan Es HH, Veldwijk M, Havenga M, Valerio D
JournalAnal Biochem
PubMed ID9177687
'A flow cytometric assay is described for the determination of glucocerebrosidase (GC) activity using fluorescein di-beta-glucopyranoside (FDGlu). Fluorescent product is formed upon intracellular hydrolysis of FDGlu and is measured in the FL1 channel of a flow cytometer. We show that the assay is specific for lysosomal beta-glucosidase or glucocerebrosidase (1) ... More
Directed evolution of a thermophilic beta-glucosidase for cellulosic bioethanol production.
AuthorsHardiman E, Gibbs M, Reeves R, Bergquist P,
JournalAppl Biochem Biotechnol
PubMed ID19834652
Characteristics that would make enzymes more desirable for industrial applications can be improved using directed evolution. We developed a directed evolution technique called random drift mutagenesis (RNDM). Mutant populations are screened and all functional mutants are collected and put forward into the next round of mutagenesis and screening. The goal ... More
Nucleic acid detection using non-radioactive labelling methods.
AuthorsMansfield ES, Worley JM, McKenzie SE, Surrey S, Rappaport E, Fortina P
JournalMol Cell Probes
PubMed ID7477006
Nucleic acid probe-based assays are now widely used in genetic research, human identification, forensics and in a broad spectrum of clinical assays in the fields of microbiology, haematology/oncology and virology. Labelled probes are used in a variety of assay formats including dot-blots, Southern blots (DNA target), Northern blots (RNA target), ... More