Required Material And Equipment Not Included In This Kit; Workflow - PromoKine PK-CA724-488HTS Instruction Manual

2. Required Material and Equipment not included in this Kit

Adherent cells

 Reaction tubes (size depends on the volume of reaction cocktail needed)
Buffered saline solution, such as PBS, D-PBS or TBS

Fixative solution (4% paraformaldehyde in PBS)

 Permeabilization solution optimised for your cell type (for example, 0.5% Triton®
X-100 in PBS, or 0.5% saponin in PBS buffer)
 Appropriate cell culture medium
1% BSA (bovine serum albumin) in PBS, pH 7.1 – 7.4

Deionized water or 18 MΩ purified water


3. Workflow

The following protocol was developed using a final EdU concentration of 10 µM and
can be adapted for any cell type. There are many factors which can influence the
labeling such as the growth medium, the density and the type of cells. To determine the
optimal concentration for your experiment, a range of EdU concentrations should be
tested for your cell type and experimental conditions.
Principally, a similar concentration to BrdU can be used for EdU as a starting point.
Heparin can be used as anticoagulant for collection, if a whole blood sample is used.
Workflow scheme for the
Optional: Treat cells with antibodies to cell surface antigens
Optional: Treat cells with antibodies to intracellular antigens
Optional: Treat cells with cell cycle stain
* At this point the sample can be stored safely
EdU-based HTS Assay
Incubate sample with EdU
Harvest cells
Fix and permeabilize cells*
Detect / Label EdU
Wash cells well with rinse buffer
Imaging and Analyse Cells
5
Instruction Manual