In Vivo Visualization of Sporozoite Invasion

Mice were anesthetized by intraperitoneal injection of a cocktail of 50 mg/kg of ketamine (Ketaset, Fort Dodge Animal Health, Overland Park, Kansas, United States), 10 mg/kg of xylazine (AnaSed, Ben Venue Laboratories, Bedford, Ohio), and 1.7 mg/kg of acepromazine (Boehringer Ingelheim Vetmedica, St. Joseph, Missouri, United States). A portion of the right abdominal skin was removed and the peritoneal cavity opened along the rib cage. The mouse was then placed on the stage of an inverted Nikon (Tokyo, Japan) Diaphot microscope equipped with a Cooke SensiCam digital camera (Cooke, Romulus, Michigan, United States). The liver was immobilized with gauze and kept moist with warm PBS. Sporozoite infection was done on the microscope stage by bite of 100−200 infected mosquitoes. Images of sporozoites entering the liver were captured using either a fluorescein isothiocyanate long-pass filter combination or a GFP/DsRed dual band filter set (Chroma Technology, Rockingham, Vermont, United States) and imported into Image-Pro Plus software (Media Cybernetics, Silver Spring, Maryland, United States). Typical exposures times were 100 ms per image for gray tone and 300 ms for RGB images. Phototoxicity was limited by reducing light transmission to 20% with a neutral density filter.

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Frevert U., Engelmann S., Zougbédé S., Stange J., Ng B., Matuschewski K., Liebes L, & Yee H. (2005). Intravital Observation of Plasmodium berghei Sporozoite Infection of the Liver. PLoS Biology, 3(6), e192.

Publication 2005

Abdominal Acepromazine Animal Bite Camera Fluorescein Infection Intraperitoneal injection Isothiocyanate Ketamine Ketaset Light Liver Microscope Mosquitoes Mouse Peritoneal cavity Phototoxicity Rib cage Silver Skin Sporozoites Transmission Xylazine

Corresponding Organization :

Other organizations : New York University, Heidelberg University

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Protocol cited in 8 other protocols

Variable analysis

independent variables

Injection of a cocktail of ketamine, xylazine, and acepromazine

dependent variables

Images of sporozoites entering the liver

control variables

Anesthetized mice
Removal of a portion of the right abdominal skin
Opening of the peritoneal cavity along the rib cage
Placement of the mouse on the stage of an inverted Nikon Diaphon microscope
Immobilization of the liver with gauze and keeping it moist with warm PBS
Sporozoite infection done on the microscope stage by the bite of 100-200 infected mosquitoes
Use of a fluorescein isothiocyanate long-pass filter combination or a GFP/DsRed dual band filter set
Importing of images into Image-Pro Plus software
Exposure times of 100 ms per image for gray tone and 300 ms for RGB images
Reduction of light transmission to 20% with a neutral density filter

controls

No positive or negative controls were explicitly mentioned in the information provided.

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