Jun 25, 2024

Public workspaceLysosomal activity DQ-Red BSA assay

DOI
dx.doi.org/10.17504/protocols.io.j8nlk8dp6l5r/v1
  • Razaul Karim1
  • 1University of Minnesota, Team Lee
Jane Balster
Open access
DOI: dx.doi.org/10.17504/protocols.io.j8nlk8dp6l5r/v1
Protocol Citation: Razaul Karim 2024. Lysosomal activity DQ-Red BSA assay. protocols.io https://dx.doi.org/10.17504/protocols.io.j8nlk8dp6l5r/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: June 07, 2024
Last Modified: June 25, 2024
Protocol Integer ID: 102397
Keywords: Lysosomal activity DQ-Red BSA assay, Cell culture, Imaging
Funders Acknowledgement:
ASAP
Grant ID: 000592
Abstract
This protocol details the Lysosomal activity DQ-Red BSA assay.
Materials
  • Reagent:

DQ-BSA Red (Thermo Fisher Scientific)

  • Coverslips:

Poly-D-Lysin coated coverslips. Corning BioCoat (Ref: 354086).
ReagentCorning® BioCoat™ Poly-D-Lysine #1 German Glass CoverslipMerckCatalog #354086
Cell Culture/Treatments:
Cell Culture/Treatments:
1h 30m
0.5x10^4/ml of cells plated on coverslips.
PFF-488 treatment.
PBS/trypsin (0.01%) wash.
Wash
DQ-BSA Red treatment:

Amount10 undetermined Duration01:30:00 at Temperature37 °C .
1h 30m
Temperature
Fix cells using 4% PFA.
Imaging:
Imaging:
Red fluorescence signal is detected only when fluorogenic substrate is hydrolyzed by lysosomal proteases.
Leica Stellaries 8 confocal laser scanning microscope.
Image analysis:

Image J. quantification of the % area of the cell covered by DQ-Red signal divided by the number of cells (DAPI stained).
Imaging