Try this RBC lysis buffer

1X  ACK (Ammonium Chloride Potassium)

            155mM NH4Cl                      8.29g

            10mM KHCO3                        1.0g

            0.1mM EDTA                        0.2 ml (500mM)

                                                Dilute to 1 liter

BD Pharm Lyse can prove very efficient. It does not contain any fixative agent, therefore, viability is least affected.

Thank you.

You can make your own ammonium cloride solution (5%). Use thid solution for washing cell pallet. It work well. I have used it for many years.

good luck

We use the following solution to lyse the RBCs:

ACK-Lysepuffer/RBC Lysis buffer (Erythrocytenlyse)

• 155mM NH4 Cl 
• 10mM KHCO3 
• 0,1mM EDTA
• à pH 7,3
• à steril filtration (0,2µM)

Depending on RBC concentration we add1-5 ml Lysis solution to the cell pellet, resuspend it  and incubate for 1-10 min. That works very fine.

I used to use Gey's solution for RBC lysis of mouse peripheral blood and splenocytes. I'm attaching simple protocols for Gey's/ACK-based RBC lysis I found on the web. This may help you to estimate the amount of lysis buffer and the incubation time.

Erythrolyse red blood cell lysis buffer from AbD Serotec works well, but it also fixes the cells, so it is no good if you want to do live cell culture afterwards.

 After suspending MNC in 2mL of 5%FBS/95%PBS, I would add 10mL of the Ammonium chloride supplied by STEM CELL Technologies. 

8 minute incubation in this solution before centrifugation, you should end up with a white pellet. 

ACK LYSIS Buffer from life tech (invitrogen) is good as i have got good results using this to separate the RBC layer from my PBMC samples

Dr. Holger,

Tried RBC lysis on human bone marrow cells using Biolegend's RBC lysis buffer as per your suggestion. Viability of cells (other than RBCs) was excellent. I have plated them on a culture dish after doing MACS. Hope that my cells are not that stressed out. Thanks for your suggestion.

Try this RBC lysis buffer

1X  ACK (Ammonium Chloride Potassium)

            155mM NH4Cl                      8.29g

            10mM KHCO3                        1.0g

            0.1mM EDTA                        0.2 ml (500mM)

                                                Dilute to 1 liter

I used RBC lysis buffer (Cat. No: 420301) from BioLegend for bone marrow samples obtained from mouse and human. The standard protocol provided in BioLegend website worked well. The viability of cells was good.

in which solvent we have to dilute the  following solutes: Mili Q water or PBS

            155mM NH4Cl                      8.29g

            10mM KHCO3                        1.0g

            0.1mM EDTA                        0.2 ml (500mM)

Solution:

- Tris-HCl 10mM ph8

- Triton X-100 1%

- Sacarosa 11%

cited in http://www.scielo.org.ar/pdf/recyt/n14/n14a01.pdf

The contents of BD FACS Lysing Solution cat. # 349202

30.0% diethylene glycol, CAS number 111-46-6,

9.99% formaldehyde, CAS number 50-00-0,

3.51% methanol, CAS number 67-56-1.

This solution is one of the most efficient

I used the following protocol inspired by BD to minimize cell loss while keeping viability high. The procedure is gentle and some RBC's may remain intact.

I use 1 Triton-X, 320 mM sucrose, 5 mM MgCl2, Tris-HCl (pH 8), double distilled water ... autoclave... and then add 1 % SDS ...OR... TES which is 10 mM Tris-HCl (pH 7.5), 1 mM EDTA (pH 7.5) and 0.1 % SDS.