Mammalian chimaeras / Anne McLaren.

  • Anne McLaren
Date:
[1976]
Catalogue details
    24/168 page 14
    14 Techniques high a percentage of successful development after transfer as did those that developed in culture without any manipulation, and foetal growth was also unaffected; on the other hand, cultured blastocysts, aggregated or not, proved less viable and gave rise to somewhat smaller foetuses than did blasto¬ cysts taken from the uterus. Mullen «fe Carter (1973) found that pronase treatment reduced the percentage of embryos developing successfully, but aggregation had no further deleterious effect. They recommend transfer of embryos to both uterine horns rather than to one only. Aggregation is possible at any stage of cleavage. Although the 8-cell stage (about 60 hours p.c.) is usually the most convenient, embryos have been used at the 2-cell (Mintz, 19620; Mulnard, 1971) and 4-cell (Hillman, Sherman & Graham, 1972) stage, or as late morulae (Mintz, 1964йг). Most workers find that embryos will no longer aggregate once blastocyst formation has begun; Stern & Wilson (1972) report aggregation of Mate morulae/early blastocysts' at 72 hours p.c., when the mean cell number would not much exceed 16. Aggregation chimaeras can be made between more than two entire embryos (e.g. Mintz, 1965a, up to 16 embryos; Hillman et al., 1972, 15 embryos), as well as between various numbers of isolated blastomeres or parts of embryos (Hillman et al., 1972). Injection chimaeras A more difficult technique, but one that offers the opportunity of studying a number of problems inaccessible to aggregation-chimaera methods, was described by Gardner (1968). The essential steps are shown in Fig. 6. After removal of the zona pellucida with pronase, donor mouse blastocysts are disaggregated in calcium- and magnesium-free saline or versene solution and one or more cells are picked up in a fine glass pipette. The recipient blastocyst is held on a suction pipette and, using micromanipulators, a triangular hole is made in the trophoblast wall by introducing, and then separating, three very fine glass needles. The donor cells are introduced into the blastocyst cavity by inserting the injection pipette through the hole. The blastocysts are transferred to the uterus of a foster-mother. Since the mice that are born are often visibly chimaeric, the donor cell or cells must become incorporated in the inner cell mass of the recipient blastocyst. An essentially similar technique, but using two rather than three micro¬ needles, has been used by Moustafa & Brinster (1972öf) to investigate the fate of heterochronic cells transferred to mouse blastocysts. Comparison with control blastocysts suggested that the procedure did not affect the blasto¬ cyst's chances of survival. When a whole extra inner cell mass is surgically transferred from one mouse blastocyst to another (Gardner, 1971), aggregation with the resident inner cell mass usually occurs, giving rise to a chimaeric embryo.
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