Application
Flow Cytometry
Sample
Human Cell (Human colorectal carcinoma)
Permeabilization
Yes - Saponin
Gating Strategy
Population gathered excluding debris
Specification
Human colorectal carcinoma
Preparation
Cell harvesting/tissue preparation method: Trypsinisation
Sample buffer: 5% BSA in PBS
Sample buffer: 5% BSA in PBS
Fixation
Formaldehyde
Other product details
Incubation time
30 minute(s) · Temperature: 20°C · Diluent: 5% BSA in PBS (v/v)
Dilution
1/200
Secondary antibody
Dilution
1/32
Name
Non-Abcam antibody was used: Anti-Mouse IgG (whole molecule)–FITC antibody
Host species: Goat
Clonality: Polyclonal
Conjugation: Fluorescein
Host species: Goat
Clonality: Polyclonal
Conjugation: Fluorescein
Additional data
Additional Notes
- LIM1215 cells grown in 6 well plates, harvested at 80% confluence using trypsin and transferred to a 96 well V bottomed plate.
- Cells were washed once with wash buffer (5% FBS in PBS).
- Pellet was resuspended in 50 µL of Fix/Perm solution and incubated for 20 minutes in the dark at room temperature.
- 70 µL of wash buffer was added before cells were centrifuged.
- Cells were washed once with wash buffer (5% FBS in PBS).
- Pellet was resuspended in 100 uL of primary antibody in wash buffer (1/200 dilution) and incubated for 30 minutes in the dark at 4oC.
- Cells were washed once with wash buffer (5% FBS in PBS).
- Pellet was resuspended in 100 µL of anti-mouse FITC in wash buffer (1/32 dilution) and incubated for 15 minutes in the dark at room temperature.
- Cells were washed once with wash buffer (5% FBS in PBS).
- Pellet was resuspended in 100 µL of wash buffer.
- Plate was read on the Accuri C6 flow cytometer.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.
MS. Jasmine Stanley-Ahmed
Verified customer
投稿 Aug 19 2016