Affinipure donkey anti mouse

Immunochemical staining was performed following standard protocols available [71 (link)]. Briefly, primary cortical neuronal cultures were fixed at 7 DIV in 4% paraformaldehyde for 10 min. Cells were incubated overnight at 4 °C in 0.1 M PBS with 5% normal horse serum and with the proper primary antibody added (see Table 1) (MAP2; 1:1000, Sigma-Aldrich, M1406). After primary antibody incubation, cultures were washed with PBS and incubated 2h at room temperature with Alexa Fluor 488-conjugated AffiniPure donkey anti-mouse (1:100; Jackson Immunoresearch Laboratories, Inc., West Grove, PA, USA). Then, coverslips mounted with Fluoromount containing DAPI onto the surface of a slide after washes with PBS. Immunofluorescence images were taken using an Olympus B×60 epifluorescence microscope, using a 20× objective.

The zona pellucida of oocytes were removed with an acidic M2 medium, and then zona-free oocytes were fixed on the glass slide using chromosome spread solution containing 0.15% Triton X-100, 1% paraformaldehyde and 3 mM dithiothreitol. Immunofluorescent staining was performed after the slides were dried for 12 h. Primary antibodies used were rabbit anti-CENPF (Abcam ab5; 1:50), mouse anti-DRP1 (Abcam ab56788; 1:50), rabbit anti-DRP1 (Abcam ab180769; 1:100), mouse anti-DRP1(BD Biosciences 611112; 1:50), rabbit anti-BUB1 (Abcam ab9000; 1:50), rabbit anti-BUBR1 (Proteintech 115042-AP; 1:100), rabbit anti-APC2 (Proteintech 13559-1-AP; 1:50), rabbit anti-MAD1 (Abcam ab175245; 1:50), rabbit anti-MAD2 (Biolegend 924601; 1:50), human anti-centromere (CREST, Antibodies Incorporated 15-234-0001; 1:100), rabbit anti-REC8 (Proteintech 10793-1-AP; 1:50), rabbit anti-SMC3 (Abcam ab128919; 1:50) and mouse anti-Myc (Thermo Fisher R950-25; 1:100). Secondary antibodies used were Alexa Fluor 647 AffiniPure Donkey Anti-Human (1:50), Rhodamine (TRITC) AffiniPure Donkey Anti-Mouse (Jackson 715-025-150; 1:100), Alexa Fluor 488 AffiniPure Donkey Anti-Mouse (1:100), Alexa Fluor 488 AffiniPure Donkey Anti-Rabbit (Jackson 711-545-152; 1:100), and Rhodamine (TRITC) AffiniPure Donkey Anti-Rabbit (1:100). DNA was stained with 10 μg/ml Hoechst 33342.