Figure 1.
Custom-designed cryoloop and cryotop using CrystalCap Copper Magnetic.
Each cap possesses a magnetic base which is fixed on the top of cryovial (*) for storage or goniometer head for SXRD analysis. Cryoloop was used for SXRD of solutions (water, TCM-199 and VS2) and cryotop for bovine COCs and morulae.
Figure 2.
The end-station of beamline 08ID-1 (A) and close-up of sample position area (B).
1) CCD- X-ray detector. 2) Hutch camera for remote monitoring system. 3) Sample camera. 4) Goniometer head. 5) CrystalCap Copper Magnetic mounted on goniometer head. 6) X-ray tube. 7) Beam stop. 8) Liquid nitrogen supply from CryoJet.
Figure 3.
The beamline setup, sample camera and cryoject control tabs of the MxDC software at the 08ID-1 beamline.
A view from sample camera shows two oocytes on cryotop and square in the middle shows the path and size (0.1×0.1 mm) of X-ray beam.
Figure 4.
Physical appearance, 2D SXRD and 1D SXRD of Water, TCM-199 and VS2 in cryoloops at 102K.
A) Appearnce of solutions through sample camera. B) Representative 2D SXRD with display of d-spacing (Å). C) Representative 1D SXRD indicating the relative intensities of peaks vs. d-spacing (Å). As d-spacing increases, the diameter of Debye's ring decreases. All peaks in water and TCM-199 corresponded to the hexagonal ice crystals except one peak (*) in TCM-199 which corresponded to NaH2PO4.H2O.
Figure 5.
Physical appearance, 2D SXRD and 1D SXRD of bovine COCs in TCM-199 (non-vitrified control) and VS2 (vitrified) on cryotops at 102K.
A) Appearance of COCs through sample camera. B) Representative 2D SXRD with display of d-spacing. C) Representative 1D SXRD indicating the relative intensities of peaks vs. d-spacing. All peaks corresponded to the hexagonal ice crystals except one peak (*) which corresponded to NaH2PO4.H2O.
Figure 6.
Ice peaks' intensities in bovine COCs in TCM-199 (non-vitrified control) or VS2 (vitrified).
Each bar represents mean±SEM (N = 5 COCs in each group). Asterisk (*) denotes the significant difference (P<0.05) in ice peak intensity between non-vitrified and vitrified COCs at a given d-spacing. Double asterisks (**) denote the presence of ice peaks in the non-vitrified COCs only.
Figure 7.
Physical appearance, 2D SXRD and 1D SXRD of bovine morulae in TCM-199 (non-vitrified control) and VS2 (vitrified) on cryotops at 102K.
A) Appearance of morula through sample camera. B) Representative 2D SXRD with display of d-spacing (Å). C) Representative 1D SXRD indicating the relative intensities of peaks vs. d-spacing (Å). All peaks corresponded to the hexagonal ice crystals except one peak (*) which corresponded to NaH2PO4.H2O.