Comet Assay 中文操作手冊 【萊恩生物科技有限公司】
§ 請先將95℃ 、37℃ 水浴 (乾浴) 打開並達到設定溫度 §
1. 收細胞，用冰PBS使其懸浮，配成 1×105 cells/mL
2. 將Lysis solution 置於4℃ 或冰上至少20分鐘
3. 將LMAgarose置於沸水5分鐘 (記得將瓶蓋旋鬆)，後移至37℃ 水浴 20分鐘以上以確保其溫度降為37℃
4. 將細胞與LMAgarose以1:10 的比例混合，立刻將75ml的混合液以tip均勻塗抹於CometSlide上
5. 將玻片置於4℃ (避光) 10分鐘，直到LMAgarose邊緣出現 0.5m m的透明環 (可將時間延長至30分鐘，以促進樣品的黏付)
6. 將玻片浸入Lysis solution，置於4℃ 或冰上30-60分鐘
7. 輕輕敲去多餘buffer，並浸入新鮮配置的Alkaline solution (pH>13)，置於室溫 (避光) 20-60分鐘
8. 輕輕敲去多餘buffer，並浸入1xTBE 5分鐘，共兩次
9. 將玻片移入水平電泳槽，電泳10分鐘，使用電壓: (兩電極間距cm/V=1)
10. 輕輕敲去TBE，將玻片浸入70%乙醇5分鐘
11. 室溫乾燥sample
12. 以 50m l SYBR Green Solution染色，置於冰箱5分鐘，輕敲去多餘染劑並使玻片乾燥完全
13. 以螢光顯微鏡透過 FITC 濾片觀察
CometAssay® Reagents | ||||
| Catalog # | Product Name | Size | ||
| 4250-050-01 | CometAssay® Lysis Solution | 2 x 500 ml | ||
| 4250-050-02 | CometAssay® LM Agarose | 15 ml | ||
| 4250-050-05 | SYBR Green | 5 μl | ||
CometAssay® Reagent Starter Kits | ||||
| Catalog # | Product Name | Size | ||
| 4250-050-K | CometAssay® Kit 50 Samples ( 25X2 well slides) | 50 samples | ||
| 4252-040-K | CometAssay® Kit, 40 Samples (2X20 well slides) | 40 samples | ||
| 4253-096-K | CometAssay® Kit 96 Samples (1X96) | 96 samples | ||
Silver Staining Kit | ||||
| Catalog # | Product Name | Size | ||
| 4251-050-K | CometAssay® Silver (CommetAssay Kit w/silver staining reagents) | 50 samples | ||
| 4254-200-K | CometAssay® Silver Staining Kit | 200 samples | ||
CometAssay® Electrophoresis Systems | ||||
| Catalog # | Product Name | Size | ||
| 4250-050-ES | CometAssay® Electrophoresis System | 1 system | ||
| 4250-050-ESK | CometAssay® 2 Well ES Unit w/ Starter Kit | 1 system, 1 starter kit | ||
| 4252-040-ESK | CometAssay® 20 Well ES Unit w/ Starter Kit | 1 system,1 starter kit | ||
| 4252-040-ESK-PS1 | CometAssay® 20 Well ES Unit w/ Starter Kit plus Power Supply for North America | 1 system, 1 starter kit, 1 power supply | ||
| 4253-096-ESK | CometAssay® 96 Well ES Unit w/ Starter Kit | 1 system, 1 stater kit | ||
| 4253-096-ESK-PS1 | CometAssay® 96 Well ES Unit w/ Starter Kit plus Power Supply for North America | 1 system, 1 starter kit, 1 power supply | ||
| 4250-050-ESK-PS1 | CometAssay® 2 Well ES Unit w/ Starter Kit and Power Supply | 1 system, 1 starter kit, 1 power supply | ||
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