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电话: 400-6721-600 Human B4GALT1/GGTB2 ELISA Kit This package insert must be read entirely before using this product. For proper performance, use the insert provided with each individual kit received. Catalog Number EK11881 C

48 T EK11882 C

96 T EK11883 C

2 *96 T EK11884 C

5 *96 T EK11885 C

10 *96 T For the quantitative determination of human Beta-1, 4-galactosyltransferase

1 (B4GALT1/GGTB2) concentration in cell culture supernates, serum and plasma. For research use only. Not for use in diagnostic procedures. Human B4GALT1/GGTB2 ELISA Kit 03/2016

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电话: 400-6721-600 Human B4GALT1/GGTB2 ELISA Kit Table of Contents Introduction Description

1 Principle of the Assay

1 Limitations of the Procedure

1 General Information Materials Provided

3 Other Supplies required

3 Storage

5 Precaution

7 Technical Hints

9 Assay Protocol Sample Collection and Storage

9 Reagent Preparation 11,13 Assay Procedure

15 Analysis Calculation of Results

17 Typical Data

17 Sensitivity

19 Precision

19 Recovery

21 Linearity

21 Calibration

21 Sample Values

23 Specificity

23 Assay Procedure Summary

25 目录 产品介绍 背景介绍

2 检测原理

2 试剂盒检测的局限

2 基本信息 试剂盒提供的材料

4 未提供的材料设备

4 贮存

6 注意事项

8 技术要点

10 检测步骤 样本的收集和贮存

10 试剂准备 12,

14 检测步骤

16 分析 结果计算

18 典型数据

18 灵敏度

20 精密度

20 回收率

22 稀释线性

22 校准

22 样本值

24 特异性

24 检测步骤概要

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电话: 400-6721-600 Human B4GALT1/GGTB2 ELISA Kit DESCREPTION Beta-1, 4-galactosyltransferase

1 (B4GALT1), one of seven beta-1, 4-galactosyltransferases, is an enzyme commonly found in the trans-Golgi complex that adds galactose to oligosaccharides. By sequence similarity, the B4GALTs form four groups: B4GALT1 and B4GALT2, B4GALT3 and B4GALT4, B4GALT5 and B4GALT6, and B4GALT7. B4GALT1 is unique among the seven enzymes because it can be expressed either as membrane associated form or secreted form. The secreted form is restricted to lactating mammary tissues where the enzyme forms a heterodimer with alpha - lactalbumin to catalyze the synthesis of lactose. The membrane form can reside either in the Golgi apparatus, where it adds galactose to N-acetylglucosamine residues, or on cell surface, where it functions as a recognition molecule during a variety of cell to cell and cell to matrix interactions, by binding to specific oligosaccharide ligands on opposing cells or in the extracellular matrix. The two enzymatic forms result from alternate transcription initiation sites and post-translational processing. Defects in B4GALT1 are the cause of congenital disorder of glycosylation type 2D. PRINCIPLE OF THE ASSAY This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for human B4GALT1 has been pre-coated onto a microplate. Standard, samples and HRP-linked detect antibody specific for B4GALT1 are pipetted into the wells and B4GALT1 present is bound by the immobilized antibody and detect antibody following incubation. After washing, substrate solution is added to the wells and color develops in proportion to the amount of B4GALT1 bound in the initial step. The color development is stopped and the intensity of the color is measured. LIMITATIONS OF THE KIT FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES. The kit should not be used beyond the expiration date on the kit label. Do not mix or substitute reagents with those from other lots or sources. If samples generate values higher than the highest standard, dilute the samples with Assay Buffer and repeat the assay. If cell culture supernate samples require large dilution, perform an intermediate dilution in culture medium. Any variation in standard dilution, operator, pipetting technique, washing techniques, incubation time or temperature, and kit age can cause variation in binding. This assay is designed to eliminate interference by factors present in biological samples. Until all factors have been tested in the ELISA immunoassay, the possibility of interference cannot be excluded. 背景介绍 β-1, 4-半乳糖基转移酶 1(B4GALT1),是7种β-1, 4-半乳糖基转移酶中的一种,通常存在于反 式高尔基复合体中,可将半乳糖转移至寡糖.根据序列相似性,B4GALTs 可分为

4 组:B4GALT1 和B4GALT2,B4GALT3 和B4GALT4,B4GALT5 和B4GALT6 以及 B4GALT7.这7种酶中, B4GALT1 是独特的,因为它可作为膜结合形式或者分泌形式表达.分泌型只存在于哺乳期乳腺组 织中,在这里,该酶和 α-乳白蛋白形成一个异源二聚体,进而催化乳糖的合成.膜结合型要么存 在于高尔基体中,在这里,该酶添加半乳糖至 N-乙酰葡糖胺残基,要么存在于细胞表面,在多种 细胞与细胞、细胞与基质的相互作用过程中,该酶通过结合到对立细胞或细胞外基质的特定寡糖 配体上,作为一个识别分子发挥作用.这两种酶形式产生于交替的转录起始位点和翻译后加工. B4GALT1 缺失会导致 2D 型糖基化的先天失常. 检测原理 本试剂盒采用双抗体夹心酶联免疫吸附检测技术. 特异性抗人 B4GALT1 单克隆抗体预包被在 高亲和力的酶标板上.酶标板孔中加入标准品、检测样本和辣根过氧化物酶标记的检测抗体,经 过孵育,样本中存在的 B4GALT1 与固相抗体和检测抗体结合.洗涤后,加入显色底物 TMB,避 光显色.颜色反应的深浅与样本中 B4GALT1 的浓度成正比.加入终止液终止反应,在450 nm 波长(参考波长

570 -

630 nm)测定吸光度值. 试剂盒检测的局限 1. 本试剂盒用于科学研究,非诊断试剂,不能用于临床诊断. 2. 请在本试剂盒标记的有效期内使用. 3. 试剂盒的试剂不能与其他批号的试剂或其他来源的试剂混合使用. 4. 如果样本值高于标准曲线最高浓度,请用检测缓冲液稀释样本,并重新检测.如果细胞培养上 清样本需要较大的稀释倍数,请用细胞培养基进行适度稀释. 5. 任何标准品稀释、操作人员、移液技术、洗涤技术、孵育温度、试剂盒保存时间的改变,都将 影响结合反应. 6. 本试剂盒在设计上去除或降低了生物学样本中的一些内源性干扰因素,并非所有可能的影响因 素都已经去除.

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电话: 400-6721-600 Human B4GALT1/GGTB2 ELISA Kit MATERIALS PROVIDED 1. B4GALT1 Microplate:

96 well polystyrene microplate (12 strips of

8 wells) coated with a monoclonal antibody against human B4GALT1. 2. B4GALT1 Standard: Recombinant human B4GALT1 in a buffered protein base with preservatives;

lyophilized. 3. B4GALT1 Detect Antibody: HRP-conjugate anti-human B4GALT1 detect antibody;

100*liquid. 4. Standard Diluent: In some, very rare cases, an insoluble precipitate of stabilizing protein has been seen in the Standard Diluent vial. This precipitate does not interfere in any way with the performance of the test and can thus be ignored. 5. Assay Buffer (10* ): PBS with 0.5 % Tween-20 and

5 % BSA. 6. Substrate: TMB (tetramethyl-benzidine). 7. Stop Solution: 0.18M sulfuric acid. 8. Wash Buffer (20* ): PBS with

1 % Tween-20. 9. Plate Covers. All reagents should be stored at

2 - 8℃. OTHER SUPPLIES REQUIRED 1. Microplate reader capable of measuring absorbance at

450 nm, with correction wavelength set at

570 nm or

630 nm. 2. Pipettes and pipette tips. 3.50 μl to

300 μl adjustable multichannel micropipette with disposable tips. 4. Multichannel micropipette reservoir. 5. Beakers, flasks, cylinders necessary for preparation of reagents. 6. Deionized or distilled water. 7. Polypropylene test tubes for dilution. 试剂盒提供的材料 未提供的材料设备 1. 能够检测

450 nm 吸光度的酶标仪,参考波长

570 nm 或630 nm. 2. 移液器及枪头 3.

50 -

300 μl 可调多道移液器及一次性枪头 4. 多道移液器加样槽 5. 准备试剂用的试管、离心管、量筒等 6. 蒸馏水或去离子水 7. 稀释用聚丙烯试管 组分 编号 EK11881 EK11882 EK11883 EK11884 EK11885 预包被酶标板 EK1188P 48T 96T 192T 480T 960T 标准品 EK1188S

1 vial

2 vial

4 vial

10 vial

20 vial 辣根过氧化物酶 标记的检测抗体 EK1188D

1 vial

1 vial

2 vial

5 vial

10 vial 标准品稀释液 EKSP10

5 ml

5 ml

10 ml

25 ml

50 ml 10* 检测缓冲液 E0310

5 ml

5 ml

10 ml

25 ml

50 ml 显色底物 TMB E0230

11 ml

11 ml

22 ml

55 ml

110 ml 终止液 E0300

11 ml

11 ml

22 ml

55 ml

110 ml 20* 洗液 E0281

50 ml

50 ml

100 ml

250 ml

500 ml 封板膜 E0200

3 3

6 15

30 4

3

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电话: 400-6721-600 Human B4GALT1/GGTB2 ELISA Kit STORAGE Store kit reagents between

2 and 8℃. Immediately after use remaining reagents should be returned to cold storage (2 to 8℃). Expiry of the kit and reagents is stated on labels. Expiry of the kit components can only be guaranteed if the components are stored properly, and if, in case of repeated use of one component, this reagent is not contaminated by the first handling. Unopened kit Store at

2 - 8℃. Opened/ Reconstituted Reagents Diluted Wash Buffer Diluted Assay Buffer Stop Solution Standard Diluent Substrate TMB Detect Antibody May be stored for up to

1 month at

2 - 8℃ Standard for up to

1 month at ≤ -20℃ in a manual defrost freezer. Discard after use. Microplate Wells Return unused wells to the foil pouch containing the desiccant pack, reseal along entire edge of zip-seal. May be stored for up to

1 month at

2 - 8℃. Provided this is within the expiration date of the kit. 贮存 试剂盒保存于

2 - 8℃,有效期标注于标签上.只有恰当保存的试剂才是有保证的.如果试剂 盒的组分需要再次使用,请确定上一次使用之后试剂没有污染. 未开封试剂盒 贮存于

2 - 8℃.请在有效期内使用. 打开的试剂盒或重组试剂1* 洗液 1* 检测缓冲液 终止液 标准品稀释液 底物 TMB 检测抗体 在2-8℃,大约可以贮存

1 个月. 标准品 置于-20℃,大约可贮存

1 个月. 使用后丢弃. 预包被酶标板 未使用的板条请放回铝箔袋,封好封口. 在2-8℃,大约可贮存

1 个月.

5 6

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电话: 400-6721-600 Human B4GALT1/GGTB2 ELISA K........