WesternLumaxLight SuperiorHRP substrate
The best choice for routine, film-imaged Western blots WesternLumaxLight Superior HRP substrate is more
sensitive than other chemiluminescent substrates, soyou can conduct shorter exposures, load less sample,
or use up to ten-times less antibody. Save time and money with WesternBright ECL.
DescriptionWesternLumaxLight Superior is a horseradish peroxidase substrate optimized for chemiluminescent Westernblots imaged using X-ray film. WesternLumaxLightECLrequiresaslittle as ten times less antibody thanother substrates,allowing you to save precious antibodies and samples. Additionally, the WesternLumaxLightsignal is long lasting,allowing multiple exposures to be carried out without substantial signal decay.
WesternLumaxLight Superior detects equal amounts of protein with up to ten times less antibody.
Duplicate slot blots containing serial dilutions of transferrin were probed with the antibody dilutions
shown,and detected with either WesternLumaxLight Superior or Pierce ECL (Thermo Scientific) accor
-ding to themanufacturers’instructions. The blots were imaged simultaneouslyon the same piece
of film.WesternLumaxLight Superior produced thesame signal with ten times less primary and five
times lesssecondary antibody.
Customerfeedback
AftertryingWesternLumaxLight Superior,weimmediatelyswitchedfromourusualbrandandnowuseitonaweeklybasis.
Itproducesastrong,long-lastingsignalwithlowbackgroundatamuchlowercost.
AssistantProfessor,BrighamYoungUniversity
WesternLumaxLight Superiorisbetterthan(theECLsubstrate)Iusedtousebecauseofitshighsensitivity.Most of my
researchinvolvesdetectinglowamountsofinducedproteins,suchasgp91phoxandHIF-1. Ihad some challenge to
detecttheseproteinswhenIusedothersubstrates;however,WesternLumaxLight Superiorhelpstodetectthesetargets.
Now,IcandetecttheseproteinsusinglesssamplewiththeWesternLumaxLight Superiorsystem.Mywholelabhas now
switchedtoWesternLumaxLight Superior.
SeniorResearchLabSpecialist,UniversityofMichigan
WehaveswitchedtotheWesternLumaxLight Superiorreagentforstraightwesternandco-IPwesternblotting. We
haveusedlessantibodiestosuitourvariousneedseasily.Bytheway,theWesternBrightreagentalsoworkedbeaut
-ifullyduringarecentdemoofacompetitor’schemiluminescentimager!Itperformedbetterthanthevendor‘sown
proprietaryreagentmadeespeciallyforthatmachine!
LabManager,UniversityofNorthCarolinaatChapelHill
ebiomall.com
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想请问下那个公司有专门的蛋白质荧光标记试剂盒出售。最好的是CY5的,我准备做三标。性价比越高越好
那些做过的前辈们指导一下。
探针
ddH2O36.25ul
buffer(瓶3)5ul
核苷酸混合液(含DIG-dUTP)(瓶2)2.5ul
dNTPStockSolution(瓶4)2.5ul
引物F1ul(10pmol/ul)
R1ul(10pmol/ul)
酶(瓶1)0.75ul
DNA1ul(100pg)
程序:35个循环,退火温度55度。
第一次通过探针标记得到了相应的PCR标记产物,亮度和对照相当,片段大小比标记的探针模板大,证明整个过程应该没有问题。
但一个月后重新用相同的体系,相同的程序,只是探针模板的量不同却什么都扩不出来。不知道问题出在哪了!
那为什么SFDA不批准CA199CEAAFP等检测试剂盒作为癌症检测的手段呢?
TUNEL的缺点是:1),操作要求高,组织样本需要固定(即使是培养细胞,也需要固定),不恰当的固定方法对实验结果影响很大,导致背景过高或者信号过弱,因此实验结果重复性不好;有的人花上半年做一个体内的TUNEL是一点都不奇怪的。2),大部分诱导凋亡的药物也引起DNA损伤,从而产生DNA断裂,易引入假阳性;3)凋亡晚期细胞基因组大量降解,导致TUNEL标记反而减少,因此此法反应的是早期凋亡比例,不能严格反应凋亡比例,属于半定量研究。
尽管有如此多缺点,但由于其是目前为数不多的能原位标记凋亡细胞的方法,因此用于组织体内凋亡研究仍然是首选。但体外细胞实验研究,很少用此法。
凋亡检测中,TUNEL并不是过时的方法,现在研究凋亡的文献仍然常见。而且相反,还比以前多一点,因为现在体内实验越来越多了,甚至线虫的凋亡研究,都用TUNEL。
凋亡研究中,的确有几种方法过时了,当然是因为有替代方案了,比如DNAladder,电镜。至于AnnexinV是不能用来和TUNEL一起评论的,前者用于细胞,而且主要是悬浮细胞,后者主要用于组织。虽然有人也做镜下的AnnexinV观察,TUNEL的细胞staining,但这都不是主流。
回到lz的原帖,的确如大家所言,做贴壁细胞,不推荐用TUNEL。如果是经典凋亡途径,只是确定比例,用最经典,最常用的subG1法即可,如果是不确定是否是凋亡,用AnnexinV,不过贴壁细胞用此法,要注意消化时间。