2019級(jí)檢驗(yàn)英語(yǔ)7ppt課件

1、About immunology immune response 免疫反應(yīng)免疫反應(yīng)antigen-antibodymonoclonal antibody (McAb)單克隆抗體單克隆抗體polyclonal antibody多克隆抗體多克隆抗體recombinant antigen 重組抗原重組抗原serial dilutions系列稀釋系列稀釋high titer高滴度高滴度enzyme-linked immunosorbent assay (ELISA )酶聯(lián)免疫吸附試驗(yàn)酶聯(lián)免疫吸附試驗(yàn)horseradish peroxidase-conjugated goat anti-rabbit I

2、gG辣根過(guò)氧化物酶標(biāo)記的羊抗兔辣根過(guò)氧化物酶標(biāo)記的羊抗兔IgGAbout immunology precipitation : double diffusion test雙擴(kuò)散試驗(yàn)雙擴(kuò)散試驗(yàn) gel immunoelectrophoresis凝膠免疫電泳凝膠免疫電泳agglutination: direct agglutination 直接凝集試驗(yàn)直接凝集試驗(yàn) indirect agglutination間接凝集試驗(yàn)間接凝集試驗(yàn) latex agglutination test; indirect hemagglutination testradioimmunoassay 放射免疫方法放射免疫

3、方法immunoblotting: PAGE 聚丙烯酰胺凝膠電泳聚丙烯酰胺凝膠電泳 polyacrylamide gel electrophoresisimmunofluorescence 免疫熒光免疫熒光 polymerase chain reaction (PCR) 聚合酶聯(lián)反應(yīng)聚合酶聯(lián)反應(yīng)restriction fragment-length polymorphism analysis (RFLP) 限制性片段多態(tài)性分限制性片段多態(tài)性分析析restriction endonuclease: BamH ; EcoR 限制性內(nèi)切酶限制性內(nèi)切酶 (5 -GAATTC-3 ) (3 -CTTAA

4、G-5 ) About molecular biologyWestern-blot 免疫印跡免疫印跡Southren-blot (DNA-DNA hybridization)Northern-blot (DNA-RNA hybridization)DNA probe DNA探針探針I(yè)mmunochromatography 免疫層析免疫層析immunogold immunogold labelling rHIV-Ag1rHIV-Ag2anti-HIV McAbELISAELISA positive control 陽(yáng)性對(duì)照陽(yáng)性對(duì)照negative control 陰性對(duì)照陰性對(duì)照blank co

5、ntrol 空白對(duì)照空白對(duì)照coating overnight at 4C 包被過(guò)夜包被過(guò)夜blocked with 封閉封閉washed with 0.1% Tween 20 in PBS the secondary antibody 二抗二抗the absorbance 吸光度吸光度pregnancy 妊娠；受孕妊娠；受孕chorionic 絨毛膜的絨毛膜的gonadotropin 促性腺激素促性腺激素latex 乳汁；膠乳乳汁；膠乳polymer 多聚物多聚物chorioepithelioma 絨毛膜上皮癌絨毛膜上皮癌hydatid 包蟲(chóng)囊包蟲(chóng)囊stir 攪拌；動(dòng)攪拌；動(dòng)vigorous

6、ly 精力旺盛的；強(qiáng)壯的精力旺盛的；強(qiáng)壯的Lesson SevenPregnancy Testing PRINCIPLE: Human chorionic gonadotropin (HCG) appears in the urine of pregnant females, and determination of its presence generally indicates a positive test for pregnancy .Lesson SevenPregnancy Testing PRINCIPLE: Utilizing an immunologic technique

7、based on the principle of indirect agglutination, tests will detect HCG at levels of 1.5 to 2.5 International Units (IU) per milliliter. Since levels as low as 1.5 IU have been reported as early as 5 days after the first missed period, negative tests should be repeated within 1 to 2 weeks.Lesson Sev

8、enPregnancy Testing PRINCIPLE: Test components consist of an antigen in the form of suspension of HCG chemically bonded to latex polymer particles and an antiserum containing antibodies to HCG. Lesson SevenPregnancy Testing Lesson SevenPregnancy Testing PRINCIPLE: When the antiserum is mixed with ur

9、ine containing a detectable level of HCG, it is neutralized, and no agglutination occurs. This is a positive test for pregnancy. Urine that contains no HCG cannot neutralize the antiserum, resulting in agglutination of the latex reagent. This indicates a negative test for pregnancy.PRINCIPLE: Patien

10、ts with chorioepithelioma or hydatid mole(葡萄胎葡萄胎) frequently excrete HCG in the urine. Therefore a positive result obtained when testing these patients does not always indicate a positive test for pregnancy. Lesson SevenPregnancy Testing REAGENTS: Available as a kit from Roche Diagnostics .PROCEDURE

11、: 1. Drop 1 drop urine in circle on glass slide . 2. Add 1 drop antiserum reagent . 3. Stir and rotate gently for 30 sec . 4. Add 1 drop antigen reagent . 5. Mix over entire circle . 6. Rotate slide vigorously for no longer than 2 min while observing for agglutination . Lesson SevenPregnancy Testing

12、 INTERPRETATION: Report as 1. Positive test: No agglutination within 2 min. HCG present in the urine of pregnant women will inhibit agglutination of latex particles in the antigen reagent, resulting in a smooth, milky appearance after rotation.Lesson SevenPregnancy Testing INTERPRETATION: Report as

13、2. Negative test: Any degree of agglutination within 2 min. Lack of HCG in the urine of nonpregnant women permits agglutination of the antigen and antiserum reagents within a 2 min period, producing easily recognizable clumping(塊塊) of the latex particles. Lesson SevenPregnancy Testing To quantitate

14、Ag-specific IgG, serial dilutions of serum prepared from tail vein blood were added to ELISA plates coated with rSRS9 or rSAG1(50ng/well). After incubation(孵育孵育) with peroxidase-coupled goat anti-mouse IgG and its substrate(底物底物), and plates were read colorimetrically at 450 nm.Sandwich ELISA One of

15、 the most useful of the immunoassays is the two antibody sandwich ELISA. This assay is used to determine the antigen concentration in unknown samples. This ELISA is fast and accurate, and if a purified antigen standard is available, the assay can determine the absolute amount of antigen in an unknow

16、n sample. The sandwich ELISA requires two antibodies that bind to epitopes(表位表位) that do not overlap on the antigen. Sandwich ELISA This can be accomplished with either two monoclonal antibodies that recognize discrete (不連續(xù)的不連續(xù)的) sites or one batch of affinity-purified polyclonal antibodies. To util

17、ize this assay, one capture antibody is purified and bound to a solid phase typically attached to the bottom of a plate well. Antigen is then added and allowed to complex with the bound antibody. Unbound products are then removed with a wash, and a labeled second detection antibody is allowed to bin

18、d to the antigen, thus completing the sandwich. Sandwich ELISAThe assay is then quantitated by measuring the amount of labeled second antibody bound to the matrix (矩陣矩陣), through the use of a colorimetric substrate. Major advantages of this technique are that the antigen does not need to be purified prior to use, and that these