- Description
- Additional
Description
Details
Description:Mouse monoclonal antibody against Thr181 phosphorylated Tau protein
Purification:Protein G affinity purified
Target Protein:Human Thr181 phosphorylated Tau
Immunogen:KLH conjugated with a short peptide [Ala-Pro-Lys-Thr(p)-Pro-Pro-Ser-Ser-Gly-Glu-Cys], corresponding to the amino acid sequence 177-187 on the phosphorylated Tau protein.
Fusion Myeloma:Sp2/0-Ag14
Specificity:This antibody reacts with the unconjugated Thr181 phosphorylated Tau peptide.
Species Reacitvity:Human, mouse
Host / Isotype: Mouse, IgG1 Kappa
Formulation:Lyophilized from a solution in 0.01M PBS pH7.2
Reconstitution:Double distilled water is recommended to adjust the final concentration to 1.00mg/mL. Avoid repeated freeze and thaw cycle.
Storage: Store at -20oC
Research Area:Alzheimer’s disease and other neurodegenerative diseases
Background:
Tau is a highly soluble microtubule associated protein found in high concentration in neuron of central nervous system. The main function of the protein is stabilizing the microtubule in nerve fibre (axon). There are six tau isoforms which are formed by alternate splicing. Hyperphosphorylation of Tau protein results in aggregation and formation of neurofibrillary tangles, which is significantly associated with cognitive impairment in Alzheimer’s disease. Tau Thr181 phosphorylation has been used as a biomarker for the differentiation of Alzheimer’s disease and other primary dementia such as dementia with Lewy bodies..
Application:
1.Indirect ELISA: The antibody reacts with phosphorylated peptide [Ala-Pro-Lys-Thr(p)-Pro-Pro-Ser-Ser-Gly-Glu-Cys] coated ELISA plate in indirect ELISA. Cross-reactivity with un-phosphorylated peptide is very low.
2. Western Blot:The antibody detected a 46kD band in mouse brain tissue lysate.
Detect the mouse brain tissue lysates using antibody (Tau-pho181 9B4) at 1:2000 (0.5 μg/mL) dilution.
References:
If research is published using this product, please inform Anogen in order to cite the reference on this datasheet. Anogen will provide one unit of product in the same category as gratitude.
Additional
Additional Information
Product Specificity | mAb anti-Thr181 Phosphorylated Tau, 9B4 |
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Application | EIA, WB |
Size | 0.1 mg |
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以FITC和PE为例来看光谱重叠。FITC单染管的荧光会部分漏到PE通道中,而这部分需要从PE通道中减掉溢漏过来的FITC荧光。那么荧光补偿该如何调节呢?l 调节电压l 检测荧光通道的自发荧光l 每个荧光通道做单染管对照检测何为单染管对照?l 细胞l 补偿微球细胞通常是流式抗体单染管对照的样本首选,但是当细胞数量有限,无多余细胞用于抗体单染或感兴趣的表面抗原表达较低,阳性分群不明显不足以用来调节补偿时,建议选择补偿微球。补偿调节时要注意:l 使用未染色的细胞做PMT的电压设置l 不要使用未染色的补偿微球设置电压eBioscience提供两种补偿微球:OneComp eBeads与UltraComp eBeads。微球大小和淋巴细胞相当,可连接各种荧光标记的抗体来调节补偿。每滴微球中都含有两群:一群是可以结合抗体的阳性群,一群是不会与抗体反应的阴性群。优点:l 使用方便,一滴即可l 使用实验抗体与微球有效结合调节补偿,无需再用CD4分群来调节l 细胞替代物,适应各种孵育时间,结果稳定流式细胞术中荧光补偿该如何调节?l 能与各种种属和亚型的抗体反应ProductSpecies CompatibilityChain RecognitionFeaturesMouse IgRat IgHamster IgRabbit IgKappa Light ChainLambda Light ChainOne DropOne VialAll cell sizesViolet LaserUltraComp eBeads 01-2222√√√√※√√√√√√OneComp eBeads 01-1111√√√√※√√√√√Competitor X Anti-Mouse Ig, k√√Competitor X Anti-Rat Ig, k√√Competitor X Anti-Rat/Hamster Ig, k√√√Competitor X Plus Anti-Mouse Ig, k√√Competitor X Plus Anti-Rat Ig, k√√Competitor Z Anti-Mouse Bead Kit√√√
竞争品牌的微球是分种属的,使用不同抗体要购买多个产品以使用注:兔来源的抗体与微球结合力较弱,但某些还是可以使用l 适用于各种激发光---适用于488 nm 蓝光,532 nm 绿光,561 nm 黄光,633-635 nm 红光,UltraComp eBeads尤其适用紫外 (355 nm) 或者 紫色 (405 nm) 激光器l 微球本身信号经过优化适合补偿调节l 价格相当,有小包装操作步骤:1. 准备好流式管,每种荧光抗体一个;2. 将微球翻转或者涡旋混匀;3. 在每管中加入一滴微球;4. 在管中分别加入相应的 1 test 抗体(不同抗体 1 test 的用量不同,具体参见说明书);5. 敲击或者涡旋混匀;6. 2-8℃暗处孵育 15-30 分钟;7. 每管加 2ml 流式染色缓冲液,400-600 x g 离心 3-5 分钟;8. 弃上清,每管加入 0.2-0.4 mL 流式染色缓冲液;9. 混匀后上机分析
细胞内的蛋白,用荧光标记的单克隆抗体识别后,用流式可以测出每个细胞的相对荧光强度。有一种特异性的微球,可以吸附一系列不同定量分子数的抗体。这种微球吸附同样的荧光抗体,可以做出荧光强度和所吸附抗体分子数量的标准曲线。然后拿细胞检测的荧光强度和这个标准曲线对比,得到细胞内所结合的抗体数。因为单克隆抗体只结合相同的抗原表位,所以可以推算出细胞内蛋白的分子数量了。
解释结果的时候要考虑到抗体的特异性和染色的效果等影响因素。
现分享于此:
LatexBeadsPhagocytosisassay
---Materials
*Fluorescencelabeledlatexbeads(1umdiameter),2.5%aqueoussUSPension
*3%BSAcontaining25mMNa2HPO4,pH6.0
*0,3%(w/v)azide
*Culturemediumcontaining5%FBS
*Distilledwater,PBS
*Bathsonication,6(12)-wellplates
---Cellcultureandtreatment
1,Inoculateplateswith7,0104cells/cm2perwell.Incubateat37℃,5%CO2for24hr,bestuntil50-70%confluenceisreached
2,Removeculturemediumandexposethecellstotestmaterial.Incubateat37℃,5%CO2for24hr.
---Preparationofcoatedlatexbeads
1,Washlatexbeadswithdistilledwaterandpelletat10,000gfor8minatRT.
2,Resuspendlatexbeadsin3%BSAcontaining25mMNa3PO4(pH6.0)andincubateatRTfor15minwithbathsonication.
*CoatingbeadsinBSAinsuresbeadsremaininamonodispersestate.
3,Washthebeadsoncewithculturemediumcontaining5%FBS.
4,Resuspendthebeadsinculturemediumatconcentration2.0%.Thisisbeadsstock.Storedindarknessat4℃.
---Assay
1,Controlsandsamples:Intactcontrol(Nostaining)1well
-Negativecontrol(azidetreated)1well
-Normalcontrol1well
-sample5wells
*Inordertodifferentiatebetweenphagocytosedbeadsandbeadsnonspecificallyadheretothecellsurface,controlcellsareexposedto0,3%(w/v)azidefor10minpriortotheadditionofcoatedbeads.Thistreatmentcompromisesmicroglialenergeticprocessesandfewbeadswereinternalizedasobservedbyfluorescentmicroscopy.Meanfluorescenceofazide-treatedmicrogliawasusedasthenegativecontrolandwassubtractedfromvaluesobtainedinexperimentalsamples.
2,Forexperimentsusing6well-plates,15μlbeadsstockin1mlculturemediumisappliedtoeachwell.Votexthebeadsstockwellandtakeout105μlandaddinto7mlculturemedium.Bathsonificatefor10minatRTindarkness.Thisisbeadsworkingsolution.
Forexperimentsusing12well-plates,6μlbeadsstockin0.4mlculturemediumisappliedtoeachwell.
3,WasheachwelltwicewithPBSandreplacewithbeadsworkingsolution,1ml/wellfor6-wellplate,0.4ml/wellfor12well-plate.Incubateinthedarkat37℃for80-120min.
4,Removebeadsworkingsolutionandwash3timeswithPBStoremoveexcessbeads.
5,LiftthecellsbyscrappingortrypsinizationandwashthecellswishPBS.
6,StainwithPI(4ug/mlfinalconcentration)andrunforFACS.
对于六分之一效率(99.9999%)ULPA过滤器,该系统挑战将产生100个粒子/ 立方英尺的下游气溶胶浓度或将在标称1.0cfm激光粒子计数器(LPC)中产生100个粒子/分钟的计数速率。
过滤器效率以及过滤器中任何相关联的泄漏可以通过LPC简单地检测。标准集团的高输出PSL气溶胶发生器取代了传统过滤器测试和无尘室认证中使用的DOP(邻苯二甲酸二辛酯)和其他油基气溶胶发生器。
目前国内的制药企业对微粒释放系统的立项开发越来越多,园子里已经就脂质体展开了很多火热的讨论,我个人也参与其中,大家交流经验,取长补短,渐渐的找到了方向,系统而固定的讨论交流是非常有好处的。
因此在此倡议建立一个微球的交流区,让大家相互交流比如:微球产业化、技术难点、实验中遇到的问题、相关理论的探讨、最新进展交流等等
以BD出品的为例,你可以购买现成的或者可以根据你要检测的不同细胞因子和厂家定制bead array。基本原理就是试剂盒含有多种微球,每一种都是两种不同荧光强度组合,所以在流式检测的时候会形成一个矩阵。每种微球上附着有可以识别一种细胞因子的抗体。和样本孵育后,再加入荧光标记的抗不同细胞因子的抗体(同样颜色)。这样,只要微球上结合有细胞因子,这个微球就会有三种荧光了。
前两种荧光来判断是哪种微球(哪种细胞因子),第三种荧光是识别抗体上的,来定量。试剂盒还会提供标准品,用来做标准曲线。
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