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容器组件
  • 产品名称:人B细胞**瘤因子2(Bcl-2)ELISA试剂盒

  • 产品型号:96T/48T
  • 产品厂商:国产进口
  • 产品价格:1
  • 折扣价格:0
  • 产品文档:
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简单介绍:
我公司提供ELISA试剂盒均为现货,质量保证价格优惠!!专业代理EBioscience、R&D、CLOUD-CLONE CORP.(CCC,美国)等品牌检测种属包含人、大鼠、小鼠、猴、鸡、绵羊等试剂盒1万余种,价格优惠,欢迎联系!!
详情介绍:
样品收集、处理及保存方法
1.  血清:使用不含热原和内**的试管,操作过程中避免任何细胞刺激,收集血液后,3000转离心10分钟将血清和红细胞迅速小心地分离。
2.  血浆:EDTA、柠檬酸盐或肝素抗凝。3000转离心30分钟取上清。
3.  细胞上清液:3000转离心10分钟去除颗粒和聚合物。
4.  组织匀浆:将组织加入适量生理盐水捣碎。3000转离心10分钟取上清。
5.  保存:如果样本收集后不及时检测,请按一次用量分装,冻存于-20℃,避免反复冻融,在室温下解冻并确保样品均匀地充分解冻。
自备物品
  • 酶标仪(450nm)
  • 高精度加样器及枪头:0.5-10uL、2-20uL、20-200uL、200-1000uL
  • 37℃恒温箱
操作注意事项
  1.   试剂盒保存在2-8℃,使用前室温平衡20分钟。从冰箱取出的浓缩洗涤液会有结晶,这属于正常现象,水浴加热使结晶完全溶解后再使用。
  2.   实验中不用的板条应立即放回自封袋中,密封(低温干燥)保存。
  3.   浓度为0的S0号标准品即可视为阴性对照或者空白;按照说明书操作时样本已经稀释5倍,*终结果乘以5才是样本实际浓度。
  4.   严格按照说明书中标明的时间、加液量及顺序进行温育操作。
  5.   所有液体组分使用前充分摇匀。
试剂盒组成
名称 96孔配置 48孔配置 备注
微孔酶标板 12孔×8条 12孔×4条
标准品 0.3mL*6管 0.3mL*6管
样本稀释液 6mL 3mL
检测抗体-HRP 10mL 5mL
20×洗涤缓冲液 25mL 15mL 按说明书进行稀释
底物A 6mL 3mL
底物B 6mL 3mL
终止液 6mL 3mL
封板膜 2张 2张
说明书 1份 1份
自封袋 1个 1个
注:标准品(S0-S5)浓度依次为:0、10、20、40、80、160 pg/mL
试剂的准备
 20×洗涤缓冲液的稀释:蒸馏水按1:20稀释,即1份的20×洗涤缓冲液加19份的蒸馏水。
洗板方法
  1.   手工洗板:甩尽孔内液体,每孔加满洗涤液,静置1min后甩尽孔内液体,在吸水纸上拍干,如此洗板5次。
  2.   自动洗板机:每孔注入洗液350μL,浸泡1min,洗板5次。
操作步骤
  1.   从室温平衡20min后的铝箔袋中取出所需板条,剩余板条用自封袋密封放回4℃。
  2.   设置标准品孔和样本孔,标准品孔各加不同浓度的标准品50μL;
  3.   样本孔先加待测样本10μL,再加样本稀释液40μL;空白孔不加。
  4.   除空白孔外,标准品孔和样本孔中每孔加入辣根过氧化物酶(HRP)标记的检测抗体100μL,用封板膜封住反应孔,37℃水浴锅或恒温箱温育60min。
  5.   弃去液体,吸水纸上拍干,每孔加满洗涤液,静置1min,甩去洗涤液,吸水纸上拍干,如此重复洗板5次(也可用洗板机洗板)。
  6.   每孔加入底物A、B各50μL,37℃避光孵育15min。
  7.   每孔加入终止液50μL,15min内,在450nm波长处测定各孔的OD值。
结果判断
 绘制标准曲线:在Excel工作表中,以标准品浓度作横坐标,对应OD值作纵坐标,绘制出标准品线性回归曲线,按曲线方程计算各样本浓度值。
 
试剂盒性能
  •  准确性:标准品线性回归与预期浓度相关系数R值,大于等于0.9900。
  •  灵敏度:*低检测浓度小于1.0 pg/mL。
  •  特异性:不与其它可溶性结构类似物交叉反应。
  •  重复性:板内、板间变异系数均小于15%。
  •  贮藏:2-8℃,避光防潮保存。
  •  有效期:6个月
免责声明
  •   试剂盒仅供研究使用,不得用于临床实验或人体实验,否则所产生的一切后果,由实验者承担,本公司概不负责。
  •   严格按照说明书操作,实验者违反说明书操作,后果由实验者承担。

Sample collection and storages
Serum - Use a serum separator tube and allow samples to clot for 30 minutes before centrifugation for 10 minutes at approximately 3000×g. Remove serum and assay immediately or aliquot and store samples at -20℃ or -80℃.Avoid repeated freeze-thaw cycles
Plasma - Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples for 30 minutes at 3000×g at 2-8℃ within 30 minutes of collection. Store samples at -20℃or -80℃. Avoid repeated freeze-thaw cycles.
Cell culture supernates and other biological fluids - Remove particulates by centrifugation and assay immediately or aliquot and store samples at -20℃or -80℃. Avoid repeated freeze-thaw cycles.
Note:  The samples shoule be centrifugated dequately and no hemolysis or granule was allowed.
Materials required but not supplied
1.  Standard microplate reader(450nm)
2.  Precision pipettes and Disposable pipette tips.
3.  37 ℃ incubator
Precautions
1.  Do not substitute reagents from one kit to another. Standard, conjugate and microplates are matched for optimal performance. Use only the reagents supplied by manufacturer.
2.  Do not remove microplate from the storage bag until needed. Unused strips should be stored at 2-8°C in their pouch with the desiccant provided.
3.  Mix all reagents before using.
Remove all kit reagents from refrigerator and allow them to reach room temperature ( 20-25°C)
Materials supplied
Name 96 determinations 48 determinations
Microelisa stripplate 12*8strips 12*4strips
Standard 0.3ml*6tubes 0.3ml*6tubes
Sample Diluent 6.0ml 3.0ml
HRP-Conjugate reagent 10.0ml 5.0ml
20X Wash solution 25ml 15ml
Chromogen Solution A 6.0ml 3.0ml
Chromogen Solution B 6.0ml 3.0ml
Stop Solution 6.0ml 3.0ml
Closure plate membrane 2 2
User manual 1 1
Sealed bags 1 1
Note: Standard (S0 → S5) concentration was followed by:0,10,20,40,80,160 pg/ml
Reagent preparation
20×wash solution:Dilute with Distilled or deionized water 1:20.
Assay procedure
1.  Prepare all reagents before starting assay procedure. It is recommended that all Standards and Samples be added in duplicate to the Microelisa Stripplate.
2.  Add standard: Set Standard wells, testing sample wells. Add standard 50μl to standard well.
3.  Add Sample: Add testing sample 10μl then add Sample Diluent 40μl to testing sample well; Blank well doesn’t add anyting.
4.  Add 100μl of HRP-conjugate reagent to each well, cover with an adhesive strip and incubate for 60 minutes at 37°C. 
5.  Aspirate each well and wash, repeating the process four times for a total of five washes. Wash by filling each well with Wash Solution (400μl) using a squirt bottle, manifold dispenser or autowasher. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Solution by aspirating or decanting. Invert the plate and blot it against clean paper towels.
6.  Add chromogen solution A 50μl and chromogen solution B 50μl to each well. Gently mix and incubate for 15 minutes at 37°C. Protect from light.
7.  Add 50μl Stop Solution to each well. The color in the wells should change from blue to yellow. If the color in the wells is green or the color change does not
appear uniform, gently tap the plate to ensure thorough mixing.
8.  Read the Optical Density (O.D.) at 450 nm using a microtiter plate reader within 15 minutes.
Calculation of results
  • This standard curve is used to determine the amount in an unknown sample. The standard curve is generated by plotting the average O.D. (450 nm) obtained for each of the six standard concentrations on the vertical (Y) axis versus the corresponding concentration on the horizontal (X) axis.
  • First, calculate the mean O.D. value for each standard and sample. All O.D. values, are subtracted by the mean value of the zero standard before result interpretation. Construct the standard curve using graph paper or statistical software.
  • To determine the amount in each sample, first locate the O.D. value on the Y-axis and extend a horizontal line to the standard curve. At the point of intersection, draw a vertical line to the X-axis and read the corresponding concentration.
  • Any variation in operator, pipetting and washing technique, incubation time or temperature, and kit age can cause variation in result. Each user should obtain their own standard curve.
  • The sensitivity by this assay is 1.0 pg/ml
  • Standard curve
 
 
Storage:  2-8℃.
validity: six months.