• 我要登录|
  • 免费注册
    |
  • 我的丁香通
    • 企业机构:
    • 成为企业机构
    • 个人用户:
    • 个人中心
  • 移动端
    移动端
丁香通 logo丁香实验_LOGO
搜实验

    大家都在搜

      大家都在搜

        0 人通过求购买到了急需的产品
        免费发布求购
        发布求购
        点赞
        收藏
        wx-share
        分享

        戊二醛固定细胞【Harvard University】

        互联网

        8656

         

        Yu-Li Wang Lab,University of massachusetts Medical School(UMASS)

        http://ylwang.umassmed.edu/protocol/cfs/glutar.htm

        Materials

         

                                           vol (for 500 ml) of stock      mg
        
                         
        
        
        

        1. 137 mM NaCl 34.25 ml 2 M

        
        

        5 mM KCl 0.83 ml 3 M

        
        

        1.1 mM Na2 HPO4 5.5 ml 100 mM

        
        

        0.4 mM KH2 PO4 2 ml 100 mM

        
        

        2 mM MgCl2 10 ml 100 mM

        
        

        2 mM EGTA 10 ml 100 mM

        
        

        5 mM PIPES 25 ml 100 mM

        
        

        5.5 mM glucose 495

        
        

        pH 6.1 (cytoskeleton buffer), warm.

        
        

         

        
        2. Glutaraldehyde, store in tightly-sealed containers at 4o C.  
        

        3. Cytoskeleton buffer with 0.3% (w/v) Triton-X 100, 0.5% glutaraldehyde, 30 ml. Triton X-100 should be pipeted slowly with a p-200 pipetman and a trimmed tip. Wipe the outside surface of the pipet tip before adding to the buffer. Pump up and down several times to rinse out the inside surface. Mix in a fixative bottle, cap tightly, and warm in a water bath.

        4. Cytoskeleton buffer with 1% glutaraldehyde, 30 ml. Mix in a fixative bottle, cap tightly, and warm in a water bath.

        5. Cytoskeleton buffer with 0.5 mg/ml NaBH4 , 30 ml. Prepare fresh before use in a fixation box. Bubbles should be visible. Store NaBH4 desiccated in a tightly-sealed container.

        6. Phosphate buffered saline, 100 ml, warm.

        7. Fixation boxes/dished.

         

        Procedure

        1. Rinse coverslip 2x with warm phosphate buffered saline.

        2. Fix 1 min in buffer 3. Agitate periodically.

        3. Rinse 2x with cytoskeleton buffer.

        4. Fix 15 min in buffer 4. Agitate periodically.

        5. Rinse 2x with cytoskeleton buffer.

        6. Treat 5 min in NaBH4. Agitate periodically.

        7. Rinse 2x with cytoskeleton buffer.

         

        ad image
        提问
        扫一扫
        丁香实验小程序二维码
        实验小助手
        丁香实验公众号二维码
        扫码领资料
        反馈
        TOP
        打开小程序