Immunolabelling techniques were used on free-floating sections, and all the washes and antibody incubations were performed in blocking solution containing 1% horse serum and 0.2% triton in phosphate buffer saline (PBST). The following primary antibodies were applied for 24 h (or 48 h for anti-cFos antibodies) at 4 °C: mouse anti-APC/CC1 Ab-7 (Millipore, OP80, 1:100), chicken anti-mCherry (Abcam, 205402, 1:1000), rat anti-Ctip2 monoclonal (Abcam, ab18465, 1:500), rabbit anti-cFos (Synaptic System 226003, 1:1000) for P9 MS/SFR pups, rabbit anti-cFos antiserum (Abcam, 190289, 1:1000) for DREADDs experiments, mouse anti-NeuN (US Biological, 1:600), rabbit anti-Olig2 (Millipore Ab9610, 1:1000), rat anti-PDGFRα (BD Bioscience, 558774, 1:600) and rabbit anti-Tbr1 (Abcam, 1:1000). The corresponding fluorescent donkey antisera secondary (Jackson ImmunoResearch, 1:200) were applied for 2 h at room temperature. Sections were subsequently mounted using the SlowFade (Molecular Probes 536939).
Ab9610
Manufactured by BD
Sourced in Japan
The AB9610 is a laboratory equipment that serves as a centrifuge. It is designed to separate the components of a liquid mixture based on their relative densities by applying centrifugal force.
Automatically generated - may contain errors
3 protocols using ab9610
1
Immunolabelling Techniques for Neural Cell Types
2
Isolation and Analysis of Mouse OPCs
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Mouse primary oligodendrocyte progenitor cells (OPCs) were isolated from mixed glial cultures derived from postnatal day 1 mice as previously described [65 (link)]. Briefly, primary mixed glial cultures were established from the forebrains of postnatal C57BL/6 mice, and grown in media containing DMEM, 2 mM Glutamax, 1 mM sodium pyruvate, 20 mM HEPES and 10% fetal bovine serum. After 10 days, the flasks were shaken overnight to remove the OPCs [66 (link)], and OPCs were seeded on 12 mm glass cover slips coated with poly-L-lysine. At 12 h post-purification cultures, BubR1 expression in OPCs was determined by NG2/BubR1 or Olig2/BubR1 double staining. The following primary antibodies were used: anti-NG2 (Millipore, Rabbit, AB5320), anti-Olig2 (Millipore, Rabbit, AB9610), and anti-BubR1 (BD, Mouse, 612503).
3
Immunohistochemistry of Myelination in Mouse Brain
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Experimental C57BL/6 mice were deeply anesthetized by 1% pentobarbital sodium (50 mg/kg, i.p.) and transcardially perfused with 0.9% sodium chloride solution followed by 4% paraformaldehyde solution (4% PFA, pH = 7.35). Brains were dissected out, post-xed in 4% PFA and gradient dehydrated in 10%, 20%, 30% (w/v) sucrose solution and were then sliced into 30 µm coronal sections with the freezing microtome (Leica CM900, Germany). The sections were blocked in blocking solution containing 10% bovine serum albumin and then incubated with primary antibody at 4℃ overnight. After rinse for three times for at least 30 minutes with 0.01 M PBS, the slices were incubated with secondary antibody for 2 hours at room temperature. After three rinses, slices were incubated with DAPI solution for 5 minutes and then followed by washout. The sections were cover-slipped with uorescent mounting medium (Sigma-Aldrich, F4680) and imaged with confocal laser microscope (Olympus FV10-ASW, Japan).
Following antibodies were used: rabbit anti-MBP (1:1000, Abcam, ab40390), rabbit anti-Olig2 (1:1000, Millipore, AB9610), rat anti-PDGFRα (1:500, BD Bioscience, 558774), mouse anti-CC1 (1:100, Millipore, OP80), donkey anti-mouse AlexaFlour™ 488 (1:500, Thermo Fisher, A21206), donkey anti-rabbit AlexaFlour™ 568 (1:500, Thermo Fisher, A10042), donkey anti-rat AlexaFlour™ 488 (1:500, Thermo Fisher, A21208).
Following antibodies were used: rabbit anti-MBP (1:1000, Abcam, ab40390), rabbit anti-Olig2 (1:1000, Millipore, AB9610), rat anti-PDGFRα (1:500, BD Bioscience, 558774), mouse anti-CC1 (1:100, Millipore, OP80), donkey anti-mouse AlexaFlour™ 488 (1:500, Thermo Fisher, A21206), donkey anti-rabbit AlexaFlour™ 568 (1:500, Thermo Fisher, A10042), donkey anti-rat AlexaFlour™ 488 (1:500, Thermo Fisher, A21208).
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