Zebrafish Xenograft Assay: A Method to Assess the Anti-cancer Efficacy of Therapeutics on Tumorigenicity in Zebrafish In Vivo

Abstract

Source: Lee, J. Y et al.,Preclinical Assessment of the Bioactivity of the Anticancer Coumarin OT48 by Spheroids, Colony Formation Assays, and Zebrafish Xenografts. J. Vis. Exp. (2018)

This video describes the zebrafish xenograft assay to assess in vivo tumor formation and study the effect of therapeutic agents.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Zebrafish Xenograft Assay

NOTE: This technical approach is visualized as a schematic (Figure 1).

1. Preparation of stock reagents
   1. Prepare 1 L of 30x Danieau’s solution: 1740 mM NaCl, 21 mM KCl, 12 mM MgSO₄•7H2O, 18 mM Ca(NO₃)₂, and 150 mM HEPES. Adjust to the pH of the buffer to 7.6 using a pH meter.
   2. Prepare 1% (50x) tricaine solution: Dissol....

Results

Figure 1: Schematic diagram of the overall process of the zebrafish xenograft assay.

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Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
A549	ATCC	CCL-185	37 C°
RPMI 1640	Lonza	30096	4 C°
FBS	Biowest	S1520-500	-20 C°
Penicillin-streptomycin	Lonza	17-602E	-20 C°
Cell culture flask T25	SPL	70025	RT
Cell culture flask T75	SPL	70075	RT
Cell culture flask T175	SPL	71175	RT
1.5ml tube	Extragene	Tube-170-C	RT
24 well plate	SPL	30024	RT
Petridish	SPL	10100	RT
PBS solution	Hyclone	SH30256.02	RT
Trypsin-EDTA	Gibco	25-300-054	4 C°
Sodium Chloride	Sigma-Aldrich	71382	RT
Potassium chloride (KCL)	Sigma-Aldrich	P9541	RT
Magnesium sulfate heptahydrate (MgSO4.7H2O)	Sigma-Aldrich	M2773	RT
Calcium nitrate tetrahydrate (Ca(NO3)2)	Sigma-Aldrich	C1396	RT
HEPES solution	Sigma-Aldrich	H0887	RT
Ethyl 3-aminobenzoate methanesulfonate (Tricaine)	Sigma-Aldrich	E10521	RT
Phenol Red solution	Sigma-Aldrich	P0290	RT
Methylcellulose	Sigma-Aldrich	M0512	RT
1-phenyl-2-thiourea (PTU)	 Sigma-Aldrich	P7629	RT
CM-Dil dye	Invitrogen	C7001	-20 C°
Glass capillary	World Precision Instruments	TW 100F-4	RT
Micropipette puller	Shutter instrument, USA	P-97	RT
Micro injector	World Precision Instruments	PV820	RT
Syringe	KOVAX	1ml	RT
Micro loader	Eppendorf	5242956003	RT
Glass slide	Marienfeld	HSU-1000612	RT
Fluorescence microscopy	Leica	DE/DM 5000B	RT