Red blood cell lysis buffer
Ultra-rapid aseptic lysis, fully preserving the target lymphocytes.
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Instruction manual- Product Description
- Composition
- Precautions
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I. Product Introduction
This product is a sterile erythrocyte lysis solution containing ammonium chloride as the active ingredient. It is optimized for processing blood or tissue samples from humans, mice, and other species; by leveraging osmotic pressure, it selectively lyses enucleated red blood cells, which can then be removed by centrifugation, while minimally affecting nucleated cells such as lymphocytes in the sample. However, this product is not suitable for lysing nucleated red blood cells, such as those found in birds.
III. Storage Conditions
Store at 4°C; stable for up to 1 year. Transport at room temperature.
IV. Instructions for Use
Additional note: The following cleavage reaction can also be carried out at room temperature.
Blood sample
1. Collect whole blood into a centrifuge tube containing an anticoagulant, centrifuge at 400–500 × g for 5 minutes, and then aspirate and discard the supernatant.
2. Add 6–10 times the volume of this reagent to the centrifuged cell pellet, gently mix with a pipette, and incubate on ice for 1–2 minutes; during lysis, gently shake the centrifuge tube as needed.
Note: Mouse blood samples can be lysed for 1–2 minutes, whereas human blood samples require 4–5 minutes.
3. After lysis, centrifuge at 400×g at 4°C for 5 minutes; the supernatant will appear red, and use a pipette to carefully aspirate and discard it.
4. Optional step: If lysis is incomplete, steps 2–3 may be repeated once. However, in most cases, a very small number of red blood cells will not affect subsequent experiments.
5. Add 5 volumes of HBSS or PBS to the sample, resuspend the cell aggregates to wash the cells. Centrifuge at 4°C at 400×g for 3 minutes, then aspirate and discard the supernatant.
Optional: Repeat the washing step once.
6. After discarding the supernatant, resuspend the cell pellet in the required solution to obtain a sample from which red blood cells have been removed.
Optional: Processing of blood samples without the washing step (this step may be omitted if the blood sample volume is very small).1. Add 10 volumes of this reagent to the whole blood, gently mix, and incubate on ice for 4–5 minutes, shaking the tube occasionally.
Note: Mouse blood can be lysed for 4–5 minutes; human peripheral blood samples can be lysed for 10 minutes. Do not exceed 15 minutes of lysis.
2. Add 20–30 mL of HBSS or PBS (volume may be adjusted according to the sample volume), and mix thoroughly. Centrifuge at 4°C at 400×g for 5 minutes; if the supernatant appears red, carefully aspirate and discard the red supernatant.
3. Optional step: If lysis is incomplete, the lysis step may be repeated once. However, in most cases, a small number of red blood cells will not affect subsequent experiments.
4. After discarding the supernatant, resuspend the cell pellet in the required solution to obtain a sample from which red blood cells have been removed.
Tissue sample
1. Digest the tissue sample to obtain a single-cell suspension, then centrifuge and discard the supernatant.
2. Add 3–5 times the volume of this reagent to the sample, mix well, and incubate on ice for 1–2 minutes, gently shaking the tube as needed. After lysis, centrifuge at 400 × g and 4°C for 5 minutes; the resulting supernatant will appear red. Carefully aspirate and discard the red supernatant.
3. Optional step: If lysis is incomplete, repeat step 2 once. However, in most cases, a very small number of red blood cells will not affect subsequent experiments.
4. Add 5 volumes of HBSS or PBS to the sample, resuspend the cell aggregates to wash the cells. Centrifuge at 400×g and 4°C for 3 minutes, then discard the supernatant.
Optional: Repeat the washing step once.
5. After discarding the supernatant, resuspend the cell pellet in the required solution to obtain a sample from which red blood cells have been removed.
Optional: Tissue sample processing without the washing step
1. Digest the tissue sample to obtain a single-cell suspension, then centrifuge and discard the supernatant.
2. Add 5 volumes of this product to the sample (e.g., add 1 mL of this product to 200 μL of cell pellet), mix thoroughly, and incubate on ice for 1–2 minutes, gently shaking the tube as needed.
3. Add 15–20 mL of PBS or HBSS (adjust the volume according to the sample size) and mix thoroughly.
4. Centrifuge at 400 × g and 4°C for 5 minutes; the supernatant will appear red—pipette out and discard the red supernatant.
5. Optional step: If lysis is incomplete, steps 2–4 may be repeated once. However, in most cases, a very small number of red blood cells will not affect subsequent assays.
6. After discarding the supernatant, resuspend the cell pellet in the required solution to obtain a sample from which red blood cells have been removed.
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II. Composition
Item number Name Specifications 100-300 Red blood cell lysis buffer 100 mL -
V. Precautions
1. This product is sterile; the treated cells can subsequently be used in other experiments conducted under sterile conditions.
2. This product is not suitable for the lysis of nucleated red blood cells (e.g., mature erythrocytes in birds).
3. During the experiment, please wear a mask and gloves.
4. This product is intended for research use only; do not use it for in vitro diagnostic or other therapeutic purposes.
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