LEUCOPERM™Permeabilisation reagent is intended for the permeabilisation of cells to enable antibodies access to intracellular structures and leaves the morphological scatter characteristics of the cells intact. Specific formulation reduces background staining and allows simultaneous addition of permeabilisation medium and fluorochrome labelled antibodies.
If both fixation and permeabilisation reagents are required, please use LEUCOPERM™product range BUF09.
Application
This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visitwww.abdserotec.com/protocols.
Application Name
Yes
No
Not Determined
Suggested Dilution
Flow Cytometry
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
Storage
BUF09CB Permeabilisation reagent should be stored and used at room temperature. DO NOT FREEZE. Do not use reagent if a precipitate forms or discolouration occurs.
Shelf Life
12 months from date of despatch.
Instructions For Use
For the detection of cell cycle antigens such as Ki-67, PCNA and BrdU, methanol modification is recommended - seeprotocol #F5.
1. Prepare cells in the appropriate manner. Adjust cell suspension to a concentration of 1 x 107cells/ml in PBS/BSA. Whole blood samples may also be used. AbD Serotec recommend the use of EDTA anti-coagulant in these circumstances, although satisfactory results may be obtained using heparin or acid-citrate dextrose.
2. Add 100ul of cell suspension to the appropriate number of test tubes.
If required, perform staining of cell surface antigens at this stage. Following staining for the recommended period, wash cells once in PBS/BSA and discard supernatant.
3. Perform fixation of cells using appropriate fixation medium.
4. Add 3ml PBS/BSA and centrifuge for 5 minutes at 300 x g. Remove supernatant.
5. Resuspend cells in 100ul of BUF09CB (Permeabilisation Reagent).
6. Immediately add recommended volume of the appropriate directly conjugated antibody. Vortex and incubate for 30 minutes at room temperature. If using an unconjugated primary antibody, wash in 3ml of PBS/BSA (as per step 5) and then repeat step 7 using an appropriate secondary antibody. There is no requirement to add further Leucoperm™.
7. Incubate for 30 minutes at room temperature.
8. Wash cells with 3ml phosphate buffered saline and centrifuge for 5 minutes at 300 x g.
9. Wash once in PBS/BSA. Remove supernatant and resuspend cells in sheath fluid for immediate analysis or resuspend cells in 0.25ml of 0.5% formaldehyde and store them at 2-8oC in the dark. Analyse fixed cells within 24 hours.
京公网安备11010802025653 版权所有：北京逸优科技有限公司
