TMB显色液(组化或膜HRP显色用)(TMB Chromogen Solution, TMB Substrate Solution or TMB Solution for Immunohistochemistry or Blotting)是采用双组分：显色A液、显色B液，双组分比单组分更加稳定，易保存。
TMB，即3, 3′,5 ,5′-Tetramethylbenzidine，是辣根过氧化物酶的常用底物。通过辣根过氧化物酶(HRP)催化TMB显色，TMB会产生可溶性蓝色产物。蓝色产物通常可以在370nm或620-650nm测定吸光度。
本产品主要适用于免疫组化或原位杂交时HRP的显色检测，以及Western、Southern、Northern等HRP的膜显色检测，也可以用于原位检测细胞或组织内源性的过氧化物酶。
产品组分表
T743339
| 试剂盒组分名称
| 2×50mL
| 2×250mL
| 储存温度
|
T743339A
| 显色A液
| 50mL
| 250mL
| 2-8℃. Store in the dark.
|
T743339B
| 显色B液
| 50mL
| 250mL
| 2-8℃. Store in the dark. |
操作步骤
1.对于免疫组化检测：
a.参考免疫组化的实验步骤，在与辣根过氧化物酶标记的抗体孵育后，用适当洗涤液摇床震荡洗涤3-5次，每次5min。
b.TMB显色液的配制：使用前5min，将TMB显色液A液和B液1:1混合，避光放置备用。
c.切片洗涤后去除洗涤液，滴加TMB显色液使其均匀覆盖组织或细胞。
d.室温避光孵育3-30min或更长时间(可在湿盒中显色过夜)，显微镜下观察染色效果，显色至预期深浅。
e.去除染色液终止显色，洗涤液摇床震荡洗涤3-5次，每次5min。
f.切片稍甩干后封片后拍照记录。如果有必要，可以使用中性红染色液进行复染后再进行拍照记录。
2.对于膜的HRP显色检测：
a.参考相应的实验步骤，在与辣根过氧化物酶标记的抗体孵育后，用适当洗涤液洗涤3-5次，每次5min。
b.TMB显色液的配制：使用前5min，将TMB显色液A液和B液1:1混合，避光放置备用。
c.膜洗涤后去除洗涤液，滴加TMB显色液使其均匀覆盖膜。
d.室温避光孵育3-30min或更长时间(可长达数小时)，显色至预期深浅。
e.拍照记录显色结果。
3.其它样品：
其它样品参考上述细胞样品或组织切片进行操作。
注意事项：
1. 本产品为显色液A应为无色或淡黄色透明液体，显色液B应为无色透明液体。均无沉淀、颗粒及絮状物。
2. 本产品仅限于专业人员的科学研究用，不得用于临床诊断或治疗，不得用于食品或药品，不得存放于普通住宅内。
3. TMB对人体有刺激性，操作时请小心，为了您的安全和健康请注意适当防护以避免直接接触人体或吸入体内。
TMB Chromogen Solution (for IHC or Blotting) is supplied as a twocomponent system: Solution A and Solution B. Compared with singlecomponent formulations, this twopart system delivers better stability and easier storage.
TMB (3,3’,5,5’-Tetramethylbenzidine) is a widely-used substrate for horseradish peroxidase (HRP). When catalyzed by HRP, TMB generates a soluble blue-colored product. The blue product can be quantified by measuring absorbance at 370 nm or 620650 nm.
This product is mainly intended for HRPmediated chromogenic detection in immunohistochemistry and insitu hybridization, as well as membranebased HRP detection including Western, Southern and Northern blotting. It can also be applied for insitu detection of endogenous peroxidase in cells or tissues.
Components
T743339 | Components | 2×50mL | 2×250mL | Storage |
T743339A | Substrate Solution A | 50mL | 250mL | 2-8℃. Store in the dark. |
T743339B | Substrate Solution B | 50mL | 250mL | 2-8℃. Store in the dark. |
Operating Procedures
1. Immunohistochemistry
a. Follow standard IHC protocols. After incubation with HRPconjugated antibody, wash on a shaker 35 times, 5 min per wash, using appropriate washing buffer.
b. Prepare working TMB solution: mix Solution A and Solution B at a 1:1 ratio 5 min before use; keep the mixture protected from light until use.
c. Remove wash buffer from sections, add prepared TMB working solution to fully cover the tissue/cell sample.
d. Incubate at room temperature in the dark for 330 min or longer (overnight incubation in a humidified chamber is permissible). Monitor staining intensity under a microscope until the desired color development is achieved.
e. Discard the chromogen solution to stop the reaction. Wash on a shaker 3-5 times, 5 min per wash with washing buffer.
f. Briefly drain slides, mount sections and capture images. If required, perform counterstaining with Neutral Red before imaging.
2. MembraneBased HRP Detection
a. Follow relevant experimental protocols. After incubation with HRPconjugated antibody, wash 3-5 times, 5 min per wash using appropriate washing buffer.
b. Prepare working TMB solution: mix Solution A and Solution B at a 1:1 ratio 5 min before use; keep protected from light until use.
c. Remove wash buffer from the membrane, apply TMB working solution to cover the membrane evenly.
d. Incubate at room temperature in the dark for 3-30 min or longer (up to several hours), until target signal intensity is obtained.
e. Capture images of the chromogenic result.
3. Other Samples
For other sample types, follow the procedures described above for cell or tissue sections.
Notes
1. Solution A should be colourless or paleyellow clear liquid; Solution B should be colourless clear liquid. Neither solution should contain precipitates, particles or flocculent matter.
2. For researchonly use by qualified laboratory personnel. Not for clinical diagnosis or therapeutic purposes. Not for food or pharmaceutical related applications. Do not store in residential premises.
3. TMB is an irritant. Handle with caution. Wear adequate personal protective equipment to avoid skin/mucous-membrane contact and inhalation.
ZJ25F0724995
危险品化学品经营许可证（带存储）