Removal Of Blastomeres Research Articles

Laser-assisted removal of necrotic blastomeres from cryopreserved embryos that were partially damaged

Objective: To examine whether the developmental potential of embryos that were partially damaged after freezing and thawing can be improved by removal of necrotic blastomeres before embryo transfer. Design: Prospective pilot study and observational clinical series. Setting: Private hospital. Patient(s): Two hundred thirty-five infertile couples undergoing frozen embryo transfer. Intervention(s): Removal of necrotic blastomeres from frozen-thawed human embryos. Main Outcome Measure(s): Pregnancy and implantation rates. Result(s): Removal of necrotic blastomeres from partially damaged frozen-thawed embryos before transfer increased rates of pregnancy (45.7% vs. 17.1%), ongoing pregnancy (40.0% vs. 11.4%) and ongoing implantation (16.2% vs. 4.3%) compared with the control group, in which necrotic blastomeres were not removed. A similarly high implantation rate (16.7%) was seen a subsequent clinical series in which necrotic blastomeres were removed from all partially damaged embryos. Conclusion(s): The viability of partially damaged frozen-thawed embryos can be improved by removal of necrotic blastomeres before embryo transfer.

The effect of the biopsy procedure and the removal of a single blastomere from day 3 human embryos on their implantation potential.

Objective: Removal of a single blastomere for preimplantation genetic diagnosis (PGD) is a well established method; however, it is not known whether the biopsy procedure has no negative influence on the implantation potential of these manipulated embryos in humans. Possible alterations of implantation potential has no immense importance for many PGD indications, however, PGD for aneuploidy screening should sum up in improved implantation rate. The aim of the present study, therefore, was to investigate the effect of the removal of a single blastomere on the implantation potential of embryos in patients who were eligible for aneuploidy screening because of their age.

Compensation for the Lost Prospective Central Nervous System by Expansion of the Prospective Epidermis in the ‘16-Cell’ Embryos of Xenopus laevis Lacking All Animal Dorsal Blastomeres

Abstract In the normal embryo of Xenopus laevis, the central nervous system (CNS) and epidermis, a pair of main ectodermal tissues, are derived mainly from the animal dorsal (AD) and animal ventral (AV) blastomeres, respectively. The defect embryo, from which all AD blastomeres have been removed at the 16-cell stage, can develop into a normally proportioned embryo, i.e., a regulated embryo, despite the striking deficiency of the prospective CNS. To compare the contribution of the AV blastomeres to the CNS and epidermis between the normal and regulated embryos, each of the AV blastomeres was labelled by a tracer injection at the 16-cell stage, and a clonal domain originating from the labelled blastomere in these ectodermal tissues was examined. By the removal of the AD blastomeres, the clonal domains of the each AV blastomeres were expanded in a dorsal direction, and covered the regions not only in the epidermis just as in normal embryos but also in the CNS extending from the anterior to the posterior end,...

The application of a 1.48 μm diode Laser system in Assisted reproduction

The recent introduction of a 1.48-μm diode laser system has brought a new stimulus to the field of laser-assisted reproduction. This objective-delivered laser can be applied in a noncontact working mode with extreme precision using conventional microscope and culture systems. A major application is the fast and efficient opening of the zona pellucida. In the field of assisted hatching, the use of this laser has enhanced the pregnancy rates for certain patient groups. Further, laser-assisted opening of the zona pellucida facilitates the removal of polar bodies or blastomeres for preimplantation genetic diagnosis. The precise openings allow for the use of blunt-ended, flame-polished aspiration needles, which greatly reduces the risk of damage to the embryo or the biopsied cells. Further modes of application of the diode laser system are the immobilisation of spermatozoa and the cryopreservation of spermazoa in empty, cell-free zona pellucida. This makes the laser technique and especially the diode laser system a versatile tool, which will be soon an essential part of assisted reproduction techniques.

Effect of Ca2+/Mg2+-free medium on the biopsy procedure for preimplantation genetic diagnosis and further development of human embryos

In this study, the use of Ca2+/Mg2+-free medium for biopsy of human embryos at the 4- to 10-cell stage on the third day of development was evaluated. When compared with control medium containing normal concentrations of Ca2+ and Mg2+ ions, the use of Ca2+/Mg2+-free medium allows an easier removal of blastomeres as illustrated by a lower rate of cell lysis as well as by a shorter time needed to perform the procedure. Subsequent embryo development to the blastocyst stage is not affected by the choice of biopsy medium, not even when embryos are exposed to the medium for 45 min. The use of Ca2+/Mg2+-free medium thus allows for an easier biopsy procedure during pre-implantation genetic diagnosis, while it does not result in a loss of developmental potential of the embryo to the blastocyst stage.

Microsurgical Fertilization and Blastomere Removal for Genetic Analysis

Figure 1. Microsurgical Fertilization and Blastomere Removal for Genetic Analysis. The oocyte and embryo are each immobilized at the tip of a holding pipette by the application of a gentle vacuum to the surrounding zona pellucida (zp). Intracytoplasmic injection (Panel A) involves perforation of the zona pellucida and oocyte plasma membrane with a glass micropipette (8 micrometers in diameter). A single sperm (arrow) is visible at the tip of the micropipette before its expulsion into the oocyte cytoplasm, and a polar body (pb) extruded into the perivitelline space during the final maturation of the oocyte is evident. Forty-five hours after . . .

Trophectoderm microbiopsy in murine blastocysts: Comparison of four methods

This study compares four trophectoderm microbiopsy techniques for removal of blastomeres from murine blastocysts: (1) aspiration, trophectoderm pipetted through the zona; (2) incision, trophectoderm excised with a microrazor; (3) slit/excision, the zona slit and herniating trophectoderm excised; and (4) hatch/excision, trophectoderm cells excised after spontaneous hatching. Murine blastocysts were comparatively biopsied using one of four methods and contrasted to zona slit and nonmicromanipulated controls. Operative cellular injury was assessed by uptake of fluorescein isothiocyanate (FITC). Postoperative embryonic viability was assessed by blastocoele reexpansion and hatching inner cell mass development and trophectoderm plating. All techniques yielded cells available for genetic analysis. The slit/excision technique and hatch/excision techniques exhibited lower operative injury and the higher postoperative viability than aspiration or incision. The slit/excision and the hatch/excision techniques, though requiring two operative steps, appear to be the least damaging of the four methods. Therefore, they should be most applicable to human blastocysts obtained either by extended culture in vitro or by uterine lavage.

Preembryo biopsy and analysis of blastomeres by in situ hybridization

We developed a method for the biopsy of preimplantation mouse embryos (preembryos) at the four- to eight-cell stage, which uses partial zona pellucida dissection. The preembryos were collected in calcium- and magnesium-free phosphate-buffered saline solution with 0.01% ethylenediaminetetraacetic acid, 0.1 mol/L sucrose, and 4 mg/ml of bovine serum albumin to facilitate removal of blastomeres. This allows entry of a fine micropipette into the perivitelline cavity with subsequent removal of a single blastomere by gentle suction. The majority of embryos (75%) from which biopsy specimens were obtained in this fashion developed to the blastocyst stage. The blastomeres obtained were mainly intact and they were fixed to glass slides. After permeabilization, in situ hybridization was performed with chromosome X- and chromosome 3-specific probes. Human unfertilized eggs and blastomeres from human polyspermic embryos also have been analyzed by in situ hybridization with chromosome specific probes. The combination of nondestructive embryo biopsy and in situ hybridization is a possible approach for preimplantation genetic diagnosis.

Allocation of cells to the inner cell mass and trophectoderm of 3/4 mouse embryos

The allocation of cells to the inner cell mass (ICM) and trophectoderm (TE) was investigated at 6-h intervals from 78 h to 102 h after hCG injection in 3/4 mouse embryos to determine the effect of removal of a single blastomere at the 4-cell stage on early differentiation. The procedures used to produce 3/4 embryos had little effect on embryo development. Embryos that had a single blastomere removed and then re-aggregated (RA embryos) had the same total number of cells as untreated (UT) embryos except at 78 h (P less than 0.05) and 102 h (P less than 0.01) post hCG where there were slightly less cells in RA embryos. Three-quarter embryos always had significantly fewer cells than RA embryos (P less than 0.001), with an average of 74% of the total cell number of RA embryos. As expected, 3/4 embryos always had significantly fewer cells in the ICM and TE compared with RA embryos (P less than 0.001). However, the ICM:TE ratio was also significantly lower in 3/4 embryos compared with RA embryos at 84, 96, and 102 h post hCG, indicating that the allocation of cells to the ICM and TE was disturbed. The ICM:TE ratio of 3/4 embryos could not be manipulated if either an early- or late-dividing blastomere was selectively biopsied at the 4-cell stage; this suggests that the known preferential contribution of an early-dividing blastomere to the ICM is not cell autonomous.

Development ofDentalium following removal of D-quadrant blastomeres at successive cleavage stages.

Each primary micromere and macromere of the D-quadrant ofDentalium was deleted, through the mesentoblast stage, to investigate the way in which the polar lobe cytoplasm exerts its influence on development.-D and -1D embryos form an apical tuft but no posttrochal structures.-2D embryos form an apical tuft and a reduced posttrochal region without a shell. -3D and -4D are externally similar to control embryos. -1d embryos and -1c embryos have an apical tuft with a reduced number of cilia. Embryos in which both 1c and 1d are deleted lack the apical tuft.-2d embryos lack shell and most other posstrochal structures. -3d and-4d embryos appear externally equivalent to controls.The polar lobe cytoplasm exerts its influence sequentially, and as inIlyanassa the maximal effect is at the third quartet stage.

The relation between normal and abnormal development of the embryo of the frog: V. As determined by the removal of the upper blastomeres of the frog's egg

The relation between normal and abnormal development of the embryo of the frog: V. As determined by the removal of the upper blastomeres of the frog's egg