Evaluation of the technique of immunosurgery for the isolation of inner cell masses from mouse blastocysts.
Inner cell masses (ICMs) immunosurgically-isolated from 31/2-day mouse blastocysts were examined for trophoblast cell contamination and developmental capacity. Blastocysts were preincubated in rabbit anti-mouse antiserum, washed thoroughly and then incubated in complement. The ICMs were then easily dissected by drawing through a fine pipette. Various experiments confirmed that the trophectoderm had been completely removed by this treatment. Firstly, the ICMs did not bind a fluorescein-conjugated antibody directed against rabbit IgG, indicating the absence of cells exposed to the rabbit antiserum during the immunosurgical procedure. Secondly, ICMs dissected from blastocysts preincubated in a suspension of melanin granules did not include any of the trophoblast cells that had phagocytosed the granules. And, thirdly, the protein synthetic profile of these ICMs was similar to microsurgically dissected ICMs, and in particular, trophoblast specific spots were absent. The developmental capacity of immunosurgically-isolated ICMs was tested by injecting them into blastocysts and transferring to the uterus of 2 1/2-day pseudopregnant recipients. Extensive chimaerism was detected in the majority of implants, 5-6 days after transfer, but only in ICM-derived tissues. This demonstrates both the lack of trophoblast cell contamination and functional viability of these ICMs.
D000818:Animals; D001666:Binding Sites, Antibody; D001755:Blastocyst / Q000166:cytology* / Q000276:immunology; D002469:Cell Separation / Q000379:methods*; D002678:Chimera; D007106:Immune Sera; D007158:Immunologic Techniques; D051379:Mice; D010587:Phagocytosis; D014327:Trophoblasts / Q000276:immunology
D007106:Immune Sera