[±×¸²3. Rapid mixing accessoryÀÇ ¸ð½Äµµ]
1. |
Ãʱâ NATAÀÇ Çü±¤ ¼¼±â¸¦ ÃøÁ¤Çϱâ À§Çؼ Syringe A¿¡ 5 uM NATA¸¦ ÁÖÀÔÇϰí, Syringe B ¿¡ ¿ë¸ÅÀÎ 0.1 M Sodium phosphate¸¦ ÁÖÀÔÇÑ´Ù. |
2. |
Çü±¤ cell¿¡ ÁÖÀ﵃ ½Ã·áÀÇ ¾çÀ» ¾à 500 uLÁ¤µµ·Î ¼³Á¤ÇÑ ÈÄ Push bar¸¦ ´·¯ ¼¿¿¡ ½Ã·á¸¦ ÁÖÀÔÇÑ´Ù. ¼³Á¤µÈ ¾ç ¸¸ÅÀÇ ½Ã·á°¡ ÁÖÀԵǸé stop syringe°¡ trigger ½ÅÈ£¸¦ ÁÖ°í Çü±¤°è°¡ Çü±¤ ÃøÁ¤À» ½ÃÀÛ ÇÑ´Ù. À§ °úÁ¤À» 6ȸ ¹Ýº¹ÇÑ´Ù. |
3. |
ÃÖÃÊ 1ȸ ÃøÁ¤ÇÑ µ¥ÀÌÅ͸¦ Á¦¿ÜÇÑ 5°³ÀÇ µ¥ÀÌÅ͸¦ Æò±ÕÇÑ´Ù. ÀÌ µ¥ÀÌÅÍ´Â Ãʱâ NATAÀÇ Çü±¤ ¼¼±â °ªÀÌ´Ù. |
4. |
Syringe B¿¡ ÁÖÀÔµÈ 0.1 M Sodium phosphat- e¸¦ ¸ðµÎ ¹èÃâ½Ã۰í 50 uM NBS¸¦ ÁÖÀÔÇÑ´Ù. Push bar¸¦ ´·¯ ½Ã·á¸¦ µ¿½Ã¿¡ ÁÖÀÔÇÑ´Ù. Çü±¤ cell¿¡ ¼³Á¤µÈ ¾çÀÇ ½Ã·á°¡ ÁÖÀԵǸé stop syringe°¡ trigger ½ÅÈ£¸¦ ÁÖ°í Çü±¤°è°¡ Çü±¤ ÃøÁ¤À» ½ÃÀÛÇÑ´Ù. ÀÌ °úÁ¤À» 6ȸ ÀÌ»ó ¹Ýº¹Çؼ ù µ¥ÀÌÅ͸¦ Á¦¿ÜÇÑ ÀÌÈÄÀÇ µ¥ÀÌÅ͸¦ Æò±ÕÇÑ´Ù. |
5. |
ÃøÁ¤µÈ µ¥ÀÌÅ͸¦ fitting programÀ» ÀÌ¿ëÇØ¼ single exponential decay fittingÇÑ´Ù. |