二抗的偶联标记

1、二抗的偶联标记一般来讲耦联到二抗上的探针主要有酶（辣根过氧化酶HRP和碱性磷酸酶AP或其衍生物APAAP ,PAP ）, 荧光基团（FITC, RRX, TR, PE）和生物素。选用哪种探针的二抗主要取决于具体的实验。对于western blot 和ELISA，最常用的二抗是酶标二抗，而细胞或组织标记实验（细胞免疫化学，组织免疫化学，流式细胞 术）中通常使用荧光基团标记的二抗，免疫组化中也可以使用辣根过氧化酶或碱性磷酸酶标记的二抗。如 果想要更大程度的放大检测信号，可以使用biotin/avidin检测系统。下面简单介绍几种二抗探针的优缺点及其适用的实验：酶：主要有两种不同的酶耦联物，HRP和

2、AP。比较而言，前者更经济、快速、稳定，而后者较前者更为 灵敏，通常情况下，HRP广泛应用于免疫组化，western blot和ELISA中，而碱性磷酸酶（AP）更适 合于固相的免疫检测实验，如ELISA和western blot. HRP标记的二抗P号产品名称反应种 属应用货期abs20008anti-Sheep IgG h+l Antibody (Rabbit) - Adsorbed HRP ConjugatedSheepELISA: 0.1 - 0.01 ug/ml IHC: 5.0 - 1.0 ug/ml WB: 0.5 - 0.05ug/ml现货abs20001goat anti-m

3、ouse IgG-HRPMouseELISA: 0.1 - 0.01 ug/ml; IHC: 5.0 - 1.0 ug/ml; WB: 0.5 - 0.05ug/ml现货abs20002goat anti-rabbit IgG-HRPRabbitELISA:0.1 - 0.01 ug/ml; IHC: 5.0 - 1.0 ug/ml; WB: 0.5 - 0.05ug/ml现货abs20031Goat Anti-Rat IgG (H + L),HRPRatIHC 1:500 - 1:5,000; ELISA 1:5,000 - 1:100,000; WB 1:10,000 - 1:200,00

4、0 1:100 - 1:800 for most applications现货abs20005Rabbit anti-Goat IgG-HRPGoatELISA: 0.1 - 0.01 ug/ml; IHC: 5.0 - 1.0 ug/ml; WB: 0.5 - 0.05ug/ml现货荧光基团：1. FITC : FITC是广泛使用的荧光基团，但其最大的缺点是淬灭快，使用抗淬灭剂可以减轻。 FITC标记的二抗货号产品名称反应种属应用货期abs20018Goat anti-Chicken IgY(IgG)-FITCChickenHisto-/Cyto-Chemistry,Flow Cytomet

5、ry,IF 1:50 - 1:200 for most applications现货abs20003Goat anti-Mouse IgG-FITCMouseIF: 10 - 2 ug/ml; IHC: 10 -2 ug/ml; WB: 0.2 - 0.05 ug/ml现货abs20012Goat anti-Mouse IgG-FITCMouseHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:50 - 1:200 for most applications现货abs20004Goat anti-Rabbit IgG-FITCRabbitIF : 1:100-

6、200现货abs20023Goat anti-Rabbit IgG-FITCRabbitHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:50 - 1:200 for most applications现货abs20006Rabbit anti-Goat IgG-FITCGoatIF: 10 - 2 ug/ml; IHC: 10 -2 ug/ml; WB: 0.2 - 0.05 ug/ml现货2. AMCA :常用在多标记实验中，如免疫荧光、流式细胞术。其缺点是，淬灭快，且由于肉眼不能很好的 检测其所激发的蓝色荧光，因而AMCA耦联的二抗更适用于检测大量存在的

7、抗原。3.花青染料（Cy2,Cy3,Cy5） : Cy2比FITC更稳定，且在表面荧光显微镜下，Cy2的荧光FITC更强。Cy3 和Cy5比大多数荧光基团更亮，更稳定，背景更弱。二者常在共聚焦显微实验中作多标记。但Cy5的缺点 是不能用表面荧光显微镜观察，因此通常用具有远红外检测器的共聚焦显微镜观察。花青染料标记的二抗货号产品名称反应种属应用abs20028Donkey anti-Goat IgG-Cy3GoatHisto-/Cyto-Chemistry,Flow Cytometry, IF 1:100 - 1:800 for most applicationsabs20015Donkey a

8、nti-Mouse IgG-Cy3MouseHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20016Donkey anti-Mouse IgG-Cy3MouseHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20022Donkey Anti-Rabbit IgG-Cy3RabbitHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:

9、800 for most applicationsabs20024Goat anti-Rabbit IgG-Cy3RabbitHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20010Goat Anti-rabbit IgG/Cy3RbIF=1:100-1000 not yet tested in other applications.optimaldilutions/concentrations should be determined by the end user.TRITC,RR

10、X,TR : TRITC经常和FITC 一起用于双标记实验中，也可以使用RRX或TR，但TR会产生较 高的背景。在使用装有氪氢灯的激光共聚焦扫描显微镜作三标记实验时，RRX尤其有用，可以和Cy2 （或 FITC）及Cy5 一起使用，因为RRX的发射波长在Cy2和Cy5中间，且和这两者的重叠很少。?：常和FITC 一起用于流式细胞术双标记实验。 PE标记的二抗货号产品名称反应种属应用abs20007Goat Anti-Mouse IgG/PE antibodymouseFlow-Cyt=1:100-1000 IF=1:100-1000 not yet tested in other applic

11、ations. optimal dilutions/concentrations should be determined by the end user.6.生物素：生物素（biotin）可以和抗生素蛋白（avidin）发生不可逆反应而紧密结合在一起。首先加入生物素标记的二抗，在加入耦联有酶或荧光集团的avidin、ExtrAvidin或streptavidin，后者能够结合于 二抗表面的多个位点上，从而极大的放大检测信号。生物素标记的二抗.货号产品名称反应种属应用abs20030Rabbit Anti-Chicken IgY PolyclonalAntibody, Biotin Conju

12、gatedChickenWB = 1:500-2000 ELISA=1:500-1000 IHC-P=1:400-800 IHC-F = 1:400-800 ICC = 1:100-500 IF = 1:100-500其他热卖二抗abs20009Donkey Anti-rabbit IgG/APC antibodyrabbitFlow-Cyt=1:100-1000 IF=1:100-1000abs20032Goat Anti-rabbit IgG/APC antibodyrabbitFlow-Cyt=1:100-1000 IF=1:100-1000abs20033Rabbit Anti-pig

13、 IgG antibodypigWB=1:1000-5000ELISA=1:2000-10000IHC-P=1:200-1000 IHC-F = 1:200-1000abs20019Donkeyanti-ChickenIgY-AlexaFluor 488ChickenHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20026Donkeyanti-GoatIgG-AlexaFluor 488GoatHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100

14、- 1:800 for most applicationsabs20027Donkeyanti-GoatIgG-AlexaFluor 647GoatHisto-/Cyto-Chemistry,Flow Cytometry, IF 1:100 - 1:800 for most applicationsabs20029Donkey Anti-Guinea PigIgG-DyLight 405Guinea PigHisto-/Cyto-Chemistry,Flow Cytometry, IF 1:100 - 1:800 for most applicationsabs20014Donkeyanti-

15、MouseIgG-AlexaFluor 488MouseHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20017Donkeyanti-MouseIgG-AlexaFluor 594MouseHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20021Donkey anti-Rabbit IgG-AlexaFluor 594RabbitHisto-/Cyto-Chemistry,Fl

16、ow Cytometry,IF 1:100 - 1:800 for most applicationsabs20020Donkeyanti-RabbitIgG-AlexaFluor 488RabbitHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20013Goat anti-Mouse IgG-AlexaFluor488MouseHisto-/Cyto-Chemistry,Flow Cytometry,IF 1:100 - 1:800 for most applicationsabs20011Goat Anti-rabbit IgG antibodyRabbit, Qu