May-Grunwald Giemsa Staining: A Method to Stain Bone Marrow Cells
Published: April 30, 2023
Abstract
Source: Mopin, A. et al. A Detailed Protocol for Characterizing the Murine C1498 Cell Line and its Associated Leukemia Mouse Model. J. Vis. Exp. (2016).
This video describes the technique of staining bone marrow cells using May- Grünwald Giemsa Staining. In the protocol, we will stain the murine myeloid leukemia cells, C149 cells, to analyze the morphology and differential counting.
Protocol
1. In vitro Characterization of the C1498 Cell Line In vitro culture of C1498 cells Prepare complete RPMI (Roswell Park Memorial Institute) 1640 medium by adding 50 ml of fetal bovine serum (FBS), 5 ml of penicillin (100 U/ml)-streptomycin (100 µg/ml), 500 µl of 50 mM β-mercaptoethanol, 5 ml of N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid (HEPES), 5 ml of non-essential amino acids and 5 mL of sodium pyruvate to 500 ml of RPMI medium. <l…
Declarações
The authors have nothing to disclose.
Materials
| C1498 cell line | ATCC | TIB 49 | |
| 2-Mercaptoethanol, Gibco | ThermoFisher Scientific | 21985 | |
| Fetal Bovine Serum (FBS) | ThermoFisher Scientific | 10270 | |
| HEPES, Gibco (1 M) | ThermoFisher Scientific | 15630 | |
| Penicillin-Streptomycin, Gibco | ThermoFisher Scientific | 15140 | |
| RPMI 1640 Medium (Gibco, GlutaMAX Supplement) | ThermoFisher Scientific | 61870 | |
| Trypan blue solution (0.4 %) | ThermoFisher Scientific | 15250-061 | concentration 1/2 |
| Chamber & filter card (EZ Cytofunnel Shandon) | Thermo Scientific | A78710003 | |
| May-Grünwald solution | RAL Diagnostics | 320070 | |
| Giemsa R solution | RAL Diagnostics | 320310 | concentration 1/20 |
| pH 6.8 buffer solution | RAL Diagnostics | 330368 | |
| Slides (Starfrost – ground edges 90) | Knittel Glass | VS1137# 077FKB | |
| Microscope Cover Glasses, 24 x 24mm | Knittel Glass | VD1 2424 Y100 | |
| Shandon Cytospin 3 Cytocentrifuge | ThermoFisher Scientific |
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May-Grunwald Giemsa Staining: A Method to Stain Bone Marrow Cells. J. Vis. Exp. (Pending Publication), e20264, doi: (2023).