Oral Delivery of Transformed Yeast Cells into the Mouse Intestinal Immune System

Abstract

Source: Hudson, L. E. et al., Transformation of Probiotic Yeast and Their Recovery from Gastrointestinal Immune Tissues Following Oral Gavage in Mice. J. Vis. Exp. (2016)

This video showcases the oral administration of transformed yeast cells into a mouse model. The transformed yeast cells are orally administered using a gavage technique, reaching the mouse's stomach. Later, the cells migrate to the small intestine, where microfold cells engulf them and facilitate their transfer to Peyer's patches and gain access to the intestinal immune system.

Protocol

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

1. UV Mutagenesis to Generate Auxotrophic Yeast Strains

1. Generate survival curves to determine needed doses of ultraviolet (UV) irradiation
   1. Prepare YPD (yeast extract peptone dextrose) media and other reagents listed in Table 1 according to standard procedures and inoculate single colonies into 5-10 ml of YPD media. Incubate cultures on a roller drum at 30 °C O/N to saturation for at least 8 hr.
   2. Determine the cell concentratio....

Results

Figure 1. Yeast colonies grown on YPD media. Example YPD plate showing viable colony forming units (CFU) of probiotic yeast after UV irradiation. Cells were serially diluted such that individual CFU can be distinguished and counted.

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
SmartSpec 3000 Spectrophotometer	BioRad	170-2501	Example of spectrophotometer for determining cell concentration and OD600 of yeast cultures
New Brunswick Roller Drum	Eppendorf	M1053-4004	Example of roller drum for yeast culture incubation
UV Stratalinker 2400	Stratagene	400075-03	Example stratalinker
Stuart Colony Counter SC6PLUS	11983044	Fisher Scientific	Plate stand with magnification records colony count upon sensing pressure from pen
Scienceware Colony Counter	F378620002	Bel-Art Scienceware	Hand held colony counter pen
Replica plating device	Fisherbrand	09-718-1	Example of replica plating stand and pads
Velveteen squares	Fisherbrand	09-718-2
L shaped sterile cell spreaders	Fisherbrand	14665230
Deoxyribonucleic acid, single stranded from salmon testes	Sigma-Aldrich	D7656-1ML	Example carrier DNA for yeast LiOAc transformation
Gavage needles	Braintree Scientific	N-PK 002	For mice 15-20 g, the suggested needle is a 22 gauge (1.25 mm ball), 1 in long, straight reusable gavage needle. For mice weighing greater than 20 g, 20 gauge or larger straight or curved gavage needles may be used
1mL sterile slip-tip disposable tuberculin syringe	Becton Dickinson	BD 309659
Blunt forceps such as Electron Microscopy Sciences 7" (178 mm) serrated tip, broad grip forceps	Electron Microscopy Sciences	77937-28	Example of blunt forceps needed for dissection
Straight and curved dissection scissors	Electron Microscopy Sciences	72966-02 and 72966-03	Examples of scissors needed for dissection