Fluorescently Labeled T-ALL Cells Isolation: A Method to Isolate T-ALL Cells from Adult Zebrafish

Published: April 30, 2023

Abstract

Source: Pruitt, M. M., et al. Isolation of the Side Population in Myc-induced T-cell Acute Lymphoblastic Leukemia in Zebrafish. J. Vis. Exp. (2017).

This protocol describes the technique of isolating fluorescently labeled T-cell acute lymphoblastic leukemia (T-ALL) tumor cells from an adult zebrafish and quantifying them using trypan blue dye.

Protocol

1. Collection of fluorescently labeled T-ALL cells from the primary tumor fish Prepare at least 20 mL of 10% heat-inactivated fetal bovine serum in 0.9x phosphate buffered saline (FBS/PBS). Add 1 mL of FBS/PBS to one vial of heparin sodium salt (300 units). Select fish bearing tumors in more than 50% of the body, or those fish having difficulty eating or swimming. Sacrifice the fish by overdosing them with tricaine or by ice water immersion. Confirm death by lack of gill m…

Divulgaciones

The authors have nothing to disclose.

Materials

Syngeneic zebrafish (CG2) See Mizgireuv et al., 2006
rag2:c-myc plasmid Langenau Laboratory
rag2:GFP plasmid constructed in de Jong Lab rag2 promoter from Langenau Laboratory
pCS2-transposase plasmid Chien Laboratory
SteREO Discovery V8 Microscope Carl Zeiss Microscopy 495015-0008-000
Tricaine methanesulfonate (MS-222) Western Chemical, Inc Fisher Scientific – NC0342409 100 g
Fetal bovine serum (FBS) HyClone Laboratories, Inc. Fisher Scientific – SH3007103 500 mL
Phosphate-Buffered Saline (PBS) Gibco by Life Technologies 1001-023 pH 7.4 (1x) – 500 mL
Heparin sodium salt Sigma-Aldrich 2106 15 – 300 unit vials
40 µm mesh filter Falcon Corning Brand 352340 Case of 50
Trypan blue Sigma Life Science T8154 20 mL – Solution (0.4%)

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Fluorescently Labeled T-ALL Cells Isolation: A Method to Isolate T-ALL Cells from Adult Zebrafish. J. Vis. Exp. (Pending Publication), e20280, doi: (2023).

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