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