Giemsa Staining for Colony Formation Assay: An In Vitro Staining Procedure to Assess Proliferative Capability of Mesenchymal Stem Cells

Published: April 30, 2023

Abstract

Source: Schlaefli, P. et al. An Enzymatic Method to Rescue Mesenchymal Stem Cells from Clotted Bone Marrow Samples. J. Vis. Exp. (2015)

In this video, we demonstrate the Giemsa staining procedure to assess the proliferative potential of mesenchymal stem cells to form viable colonies. Giemsa stain is a neutral stain primarily composed of azure and eosin dyes, which carry selective affinity for cellular components depending on their charge.

Protocol

1. Seeding for Expansion Cell Culture and CFU Plates Resuspend the cells (consisting of erythrocytes and mononuclear cells) in 50 ml of basal medium: Alpha-MEM supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin, 100 mg/ml streptomycin and 2.5 mg/ml amphotericin B. Count the cells diluted 1:10 in Trypan Blue solution using a Neubauer chamber. Plate the cells in cell culture flasks at the density of 5 x 10⁷ cells/cm² and incubate in a humidified incubator a…

Declarações

The authors have nothing to disclose.

Materials

Basal Medium Components
PenStrep 100X Gibco 15140122
Human FGF-basic Peprotech 100-18B
MEM Alpha w/ Nucleoside, w/ stable Glutamine Amimed 1-23S50-I
FBS Heat Inactivated Amimed 2-01F36-I
Amphotericin B Applichem A1907
Generic
PBS Applichem 964.91
Giemsa stain Applichem A0885
Consumables
Primaria cell cuture dish Falcon 353803
Cell culture flask – Flask T300 TPP 90301
Equipment
Stripettes Serological Pipette 5ml Corning 4487-200ea
Humidified incubator Heracells 240 Thermo scientific 51026331
Heraeus Multifuge 1S-R Thermo scientific 75004331

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Giemsa Staining for Colony Formation Assay: An In Vitro Staining Procedure to Assess Proliferative Capability of Mesenchymal Stem Cells. J. Vis. Exp. (Pending Publication), e21050, doi: (2023).

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