Red Blood Cell Lysis Buffer

Red Blood Cell Lysis Buffer (ACK Lysis Buffer) is a classic lysis solution designed to remove red blood cells without damaging nucleated cells. The erythrocyte lysate lyses tissue cells that do not contain erythrocytes and can be further used for primary culture, cell fusion, flow cytometric analysis, separation and extraction of nucleic acids and proteins. The basic principle is to lyse erythrocytes by using the difference in intracellular osmolarity to cause the cell membrane to swell. The main components include ammonium chloride, potassium bicarbonate and EDTA-Na2. Ammonium ions cannot pass through the cell membrane, while other ions can, resulting in the difference in ion concentration between the inside and outside of the cell, and the diffusion of external water into the cell, causing the red blood cells to swell and achieve the lysis effect. The product is filtered and sterilized by 0.2 μm filter membrane.

Red Blood Cell Lysis Buffer should be shipped with wet ice; Store at 2-8℃, valid for 12 months.

Tissue/cell samples:

1. Fresh tissue is digested by trypsin or collagenase and dispersed into individual cell suspension. Centrifuge at 300-400 g for 5 min at 4°C and discard the supernatant.

2. Add red blood cell lysis buffer to the cell precipitation at a ratio of 1:3-5, mix with gently blowing and lyse for 1-2 min.

3. Centrifuge at 800-1000 rpm for 5-8 min at 4°C, then discard the red supernatant. Repeat steps 2 and 3 if lysis is incomplete.

4. The cells were resuspended by adding 3-5 mL of Hank's solution or serum-free culture medium to the precipitated fraction, and then centrifuged at 300-400 g for 3-5 min at 4°C. This step was repeated 2-3 times to wash the cells.

5. Resuspended cells with appropriate solution as required for subsequent experiments; For RNA extraction, it is preferable to use the solution prepared with DEPC water at the beginning of step 4.

1. Fresh anticoagulant blood, discard the supernatant after centrifugation.

2. Pre-estimate the volume of cell precipitation and add 2-8°C pre-cooled red blood cell lysis buffer into the cell precipitation at a ratio of 1:6-10. For example, for 0.2 mL of cell precipitation, add 1.2-2.0 mL of red blood cell lysis buffer, gently blow to mix, and lysed for 1-5 min at 4°C or room temperature.

3. Centrifuge at 800-1000 rpm for 5-8 min at 4°C, then discard the red supernatant. If erythrocyte lysis is incomplete, repeat steps 2 and 3 once;

4. The cells were resuspended by adding Hank's solution or serum-free culture medium to the precipitated fraction, and then centrifuged at 300-400 g for 3-5 min at 4°C. This step was repeated 2-3 times to wash the cells;

5. Resuspended cells with appropriate solution as required for subsequent experiments; For RNA extraction, it is preferable to use the solution prepared with DEPC water at the beginning of step 4.

1. This product has been filtered and sterilized. Attention should be paid to aseptic operation to avoid contamination.

2. If the follow-up test is used for cell culture, the operation should be completed in an ultra-clean bench with attention to aseptic operation to avoid contamination of the cells and affecting the cell culture.

3. After centrifugal washing, if a very small amount of erythrocytes are found, the test can be continued without affecting the subsequent detection.

4. For your safety and health, please wear safety glasses, gloves, or protective clothing.

For Research Use Only!