Human Erythrocyte Lysing Kit

R&D Systems | Catalog # WL1000

R&D Systems
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Key Product Details

Species

Human

Product Summary for Human Erythrocyte Lysing Kit

Kit Summary

For the removal of erythrocytes (red blood cells) from whole blood.

Key Benefits

  • Gently lyses erythrocytes
  • Stabilizes leukocytes for analysis by flow cytometry
  • Maintains natural light scattering and fluorescent staining properties of leukocytes
  • Maintains leukocyte viability for subsequent cell culture manipulation
 

Why Remove Erythrocytes?

The lysis of erythrocytes from whole blood is an important initial step in the isolation and analysis of enriched leukocyte preparations. Recovered immune cells can be accurately characterized following red blood cell removal. Lysis of erythrocytes under conditions that do not disrupt lymphocytes or myeloid cells is critical for downstream applications utilizing leukocytes harvested from whole blood.

Reagents Provided

The Human Erythrocyte Lysing Kit contains the following reagents for the controlled lysis of erythrocytes in whole blood.

  • H-Lyse Buffer Concentrate (10X)
  • Wash Buffer Concentrate (10X)
  • Fixative Concentrate (10X)

*This kit contains sufficient reagents to process 250 mL of whole blood.

Stability and Storage

Store all reagents at 20 °C to 25 °C. 

 

Data Examples


View Larger Image

Lysis of Erythrocytes in Whole Blood. Forward light scatter (FSC-H) vs. side light scatter (SSC-H) dot histograms of human blood before (A) and after (B) treatment with the Human Erythrocyte Lysing Kit. (C) Dot histogram of whole blood stained with PE-conjugated Mouse Anti-Human CD14 Monoclonal Antibody (Catalog # FAB3832P) and APC-conjugated Mouse Anti-Human CD3 epsilon Monoclonal Antibody (Catalog # FAB100A) followed by erythrocyte lysis using the Human Erythrocyte Lysing Kit. Stained cells were stored for 24 hours at 2 °C to 8 °C prior to flow cytometric analysis.

Formulation, Preparation, and Storage

Shipping

The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.

Storage

Store the unopened product at room temperature. Do not use past expiration date.

Product Documents for Human Erythrocyte Lysing Kit

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

Product Specific Notices for Human Erythrocyte Lysing Kit

For research use only

⚠ WARNING: This product can expose you to chemicals including formaldehyde and methanol, which are known to the State of California to cause cancer and reproductive toxicity with developmental effects. For more information, go to www.P65Warnings.ca.gov.

Citations for Human Erythrocyte Lysing Kit

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Protocols

View specific protocols for Human Erythrocyte Lysing Kit (WL1000):

Refer to the product datasheet for complete product details.

Briefly, erythrocytes can be lysed in whole blood using the following procedure:

  • Stain whole blood or an enriched mononuclear cell preparation with antibody
  • Add H-Lyse Buffer
  • Wash cells with Wash Buffer
  • Fix cells for flow cytometry with Fixative
 
 

Reagents Provided

The Human Erythrocyte Lysing Kit contains the following reagents for the controlled lysis of erythrocytes in whole blood:

  • H-Lyse Buffer Concentrate (10X)
  • Wash Buffer Concentrate (10X)
  • Fixative Concentrate (10X)

This kit contains sufficient reagents to process 250 mL of whole blood.

Other Supplies Required

  • Ficoll-Hypaque™
  • Sterile distilled or deionized water
  • Sterile centrifuge tubes
  • Benchtop centrifuge
  • Pipettes and sterile pipette tips

Procedure Overview

R&D Systems Protocol for the Lysis of Erythrocytes Using the Mouse Erythrocyte Lysing Kit

 

 
 
 
 

If the cell preparation will be further processed with a T cell enrichment column, complete these steps prior to treating cells with H-Lyse Buffer.

 
  1. Stain 100 uL of whole blood with antibody or antibodies (if performing flow cytometry).
  2. Vortex cell pellet vigorously.
  3. Add 2 mL 1X H-Lyse Buffer.
  4. Vortex vigorously.
  5. Incubate the cells for 5-10 minutes at room temperature.
  6. Pellet leukocytes using centrifugation.
  7. Wash the pelleted leukocytes with 2 mL 1X Wash Buffer.
  8. Resuspend the cells in 1 mL 1X Wash Buffer.
  9. Fix the cells with 100 uL 10X Fixative Concentrate if flow cytometry analysis will be delayed more than one hour.
  10. Or
  11. Utilize leukocytes for alternate downstream applications
  12. Prepare a single cell suspension of mononuclear cells.
  13. Wash the cells with excess sterile PBS.

Technical Hints

  • If flow cytometric analysis of the cells will be delayed for more than 1 hour following erythrocyte lysis, the cells can be fixed at this time to stabilize them for later analysis. This step should be eliminated if cells are to be used for cell culture.
  • Fixed cells should be stored at 2 °C to 8 °C until flow cytometric analysis. Although stained cells will be stable for up to 48 hours, we recommend that flow cytometric analysis be performed as soon as possible.

FAQs

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