人B-淋巴细胞趋化因子1(BLC-1;CXCL13)ELIS

本试剂盒只能用于科学研究，不得用于医学诊断 本试剂盒只能用于科学研究，不得用于医学诊断 本试剂盒只能用于科学研究，不得用于医学诊断 本试剂盒只能用于科学研究，不得用于医学诊断

人人人人（（（（Human Human Human Human））））B-B-B-B-淋巴细胞趋化因子 淋巴细胞趋化因子 淋巴细胞趋化因子 淋巴细胞趋化因子 1111（（（（BLC-1/CXCL13 BLC-1/CXCL13 BLC-1/CXCL13 BLC-1/CXCL13））））
ELISA ELISA ELISA ELISA 检测试剂盒 检测试剂盒 检测试剂盒 检测试剂盒使用说明书检测原理 检测原理 检测原理 检测原理试剂盒采用双抗体一步夹心法酶联免疫吸附试验（ELISA）。往预
先包被B-淋巴细胞趋化因子1（BLC-1/CXCL13）抗体的包被微孔中，
依次加入标本、标准品、HRP标记的检测抗体，经过温育并彻底洗涤。
   
用底物TMB显色，TMB在过氧化物酶的催化下转化成蓝色，并在酸的
作用下转化成最终的黄色。颜色的深浅和样品中的B-淋巴细胞趋化
因子1（BLC-1/CXCL13）呈正相关。用酶标仪在450nm 波长下测定吸

光度（OD 值），计算样品浓度。
样品收集、处理及保存方法 样品收集、处理及保存方法 样品收集、处理及保存方法 样品收集、处理及保存方法
1. 血清：使用不含热原和内毒素的试管，操作过程中避免任何细胞
刺激，收集血液后，3000 转离心 10 分钟将血清和红细胞迅速小心地
分离。2. 血浆：EDTA、柠檬酸盐或肝素抗凝。3000 转离心 30 分钟取上清。
3. 细胞上清液：3000 转离心 10 分钟去除颗粒和聚合物。
4. 组织匀浆：将组织加入适量生理盐水捣碎。3000 转离心 10 分钟
取上清。5. 保存：如果样本收集后不及时检测，请按一次用量分装，冻存于
-20℃，避免反复冻融，在室温下解冻并确保样品均匀地充分解冻。
自备物品 自备物品 自备物品 自备物品1. 酶标仪（450nm）2. 高精度加样器及枪头：0.5-10uL、2-20uL、20-200uL、200-1000uL
3. 37℃恒温箱操作注意事项 操作注意事项 操作注意事项 操作注意事项1. 试剂盒保存在 2-8℃，使用前室温平衡 20 分钟。从冰箱取出的
浓缩洗涤液会有结晶，这属于正常现象，水浴加热使结晶完全溶解
后再使用。2. 实验中不用的板条应立即放回自封袋中，密封（低温干燥）保存。
 3. 浓度为 0 的 S0 号标准品即可视为阴性对照或者空白；按照说明
   
书操作时样本已经稀释 5 倍，最终结果乘以 5 才是样本实际浓度。
4. 严格按照说明书中标明的时间、加液量及顺序进行温育操作
5. 所有液体组分使用前充分摇匀。
试剂盒组成 试剂盒组成 试剂盒组成 试剂盒组成名称名称名称名称 96969696 孔配置 孔配置 孔配置 孔配置      48484848 孔配置 孔配置 孔配置 孔配置 
备注备注备注备注微孔酶标板 12 孔×8 条   12 孔×4 条 
无标准品 0.3mL*6 管  0.3mL*6 管 
无样本稀释液 6mL 3mL 无检测抗体-HR 10mL 5mL 无20×洗涤缓冲液 25mL 15mL 按说明书进行稀释底物 A 6mL 3mL 无底物 B 6mL 3mL 无终止液 6mL 3mL 无封板膜 2 张 2 张 无说明书 1 份 1 份 无自封袋 1 个 1 个 无注：标准品（S0-S5）浓度依次为：0、0.5、1、2、4、8 ng/mL
试剂的准备 试剂的准备 试剂的准备 试剂的准备20×洗涤缓冲液的稀释：蒸馏水按 1：20 稀释，即 1 份的 20×洗涤缓
冲液加 19 份的蒸馏水。
洗板方法 洗板方法 洗板方法 洗板方法1. 手工洗板：甩尽孔内液体，每孔加满洗涤液，静置 1min 后甩尽
孔内液体，在吸水纸上拍干，如此洗板 5 次。
2. 自动洗板机：每孔注入洗液 350μL，浸泡 1min，洗板 5 次。
操作步骤 操作步骤 操作步骤 操作步骤1. 从室温平衡 20min 后的铝箔袋中取出所需板条，剩余板条用自封
袋密封放回 4℃。2. 设置标准品孔和样本孔，标准品孔各加不同浓度的标准品 50μL；
3. 样本孔先加待测样本 10μL，再加样本稀释液 40μL；空白孔不
加。4. 除空白孔外，标准品孔和样本孔中每孔加入辣根过氧化物酶
（HRP）标记的检测抗体 100μL，用封板膜封住反应孔，37℃水浴
锅或恒温箱温育 60min。
5. 弃去液体，吸水纸上拍干，每孔加满洗涤液，静置 1min，甩去洗涤液，吸水纸上拍干，如此重复洗板 5 次（也可用洗板机洗板）。
   
6. 每孔加入底物 A、B 各 50μL，37℃避光孵育 15min。
7. 每孔加入终止液 50μL，15min 内，在 450nm 波长处测定各孔的

OD 值。结果判断 结果判断 结果判断 结果判断绘制标准曲线：在 Excel 工作表中，以标准品浓度作横坐标，对应
OD 值作纵坐标，绘制出标准品线性回归曲线，按曲线方程计算各样
本浓度值。试剂盒性能 试剂盒性能 试剂盒性能 试剂盒性能1. 准确性：标准品线性回归与预期浓度相关系数 R 值，大于等于
0.9900。2. 灵敏度：最低检测浓度小于 0.1 ng/mL。
3. 特异性：不与其它可溶性结构类似物交叉反应。
4. 重复性：板内、板间变异系数均小于 15%。
5. 贮藏：2-8℃，避光防潮保存。
6. 有效期：6 个月免责声明 免责声明 免责声明 免责声明1. 试剂盒仅供研究使用，不得用于临床实验或人体实验，否则所
产生的一切后果，由实验者承担，本公司概不负责。
2. 严格按照说明书操作，实验者违反说明书操作，后果由实验者
承担。FOR FOR FOR FOR RESEARCH RESEARCH RESEARCH RESEARCH USE USE USE USE ONLY. ONLY. ONLY. ONLY.
NOT NOT NOT NOT FOR FOR FOR FOR USE USE USE USE ININININ DIAGNOSTIC DIAGNOSTIC DIAGNOSTIC DIAGNOSTIC PROCEDURES PROCEDURES PROCEDURES PROCEDURES....
Human Human Human Human BLC-1/CXCL13 BLC-1/CXCL13 BLC-1/CXCL13 BLC-1/CXCL13 ELISA ELISA ELISA ELISA Kit Kit Kit Kit instruction instruction instruction instruction
Intended Intended Intended Intended use use use use
This BLC-1/CXCL13 ELISA kit is intended Laboratory for Research use only and
is not for use in diagnostic or therapeutic procedures.The Stop Solution changes th
color from blue to yellow and the intensity of the color is measured at 450 nm using
a spectrophotometer. In order to measure the concentration of BLC-1/CXCL13 in
the sample, this BLC-1/CXCL13 ELISA Kit includes a set of calibration standards.
The calibration standards are assayed at the same time as the samples and allow the
operator to produce a standard curve of Optical Density versus BLC-1/CXCL13
concentration. The concentration of BLC-1/CXCL13 in the samples is then
determined by comparing the O.D. of the samples to the standard curve.
SSSSample ample ample ample collection collection collection collection and and and and storages storages storages storages
Serum Serum Serum Serum - Use a serum separator tube and allow samples to clot for 30 minutes
before centrifugation for 10 minutes at approximately 3000×g. Remove serum and
assay immediately or aliquot and store samples at -20℃ or -80℃.Avoid repeated
freeze-thaw cycles
Plasma Plasma Plasma Plasma - Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge
samples for 30 minutes at 3000×g at 2-8℃ within 30 minutes of collection. Store
samples at -20℃or -80℃. Avoid repeated freeze-thaw cycles.
Cell Cell Cell Cell culture culture culture culture supernates supernates supernates supernates and and and and other other other other biological biological biological biological fluids fluids fluids fluids ---- Remove particulates
by centrifugation and assay immediately or aliquot and store samples at -20℃or
-80℃. Avoid repeated freeze-thaw cycles.
Note: Note: Note: Note:  The samples shoule be centrifugated dequately and no hemolysis or
granule was allowed.
Materials Materials Materials Materials required required required required but but but but not not not not supplied supplied supplied supplied
1.  Standard microplate reader(450nm)
2. Precision pipettes and Disposable pipette tips.
3. 37 ℃ incubator
PPPPrecautions recautions recautions recautions
1. Do not substitute reagents from one kit to another. Standard, conjugate and
microplates are matched for optimal performance. Use only the reagents supplied by
manufacturer.
2. Do not remove microplate from the storage bag until needed. Unused strips
should be stored at 2-8°C in their pouch with the desiccant provided.
3. Mix all reagents before using.
Remove all kit reagents from refrigerator and allow them to reach room temperature( 20-25°C)
Materials Materials Materials Materials supplied supplied supplied supplied
Name 96 determinations  48 determinations
Microelisa stripplate 12*8strips 12*4strips
Standard 0.3ml*6tubes 0.3ml*6tubes
Sample Diluent 6.0ml 3.0mlHRP-Conjugate reagent 10.0ml 5.0ml20X Wash solution 25ml 15mlChromogen Solution A 6.0ml 3.0mlChromogen Solution B 6.0ml 3.0mlStop Solution 6.0ml 3.0mlClosure plate membrane 2 2User manual 1 1Sealed bags 1 1Note: Note: Note: Note: Standard (S0 → S5) concentration was followed by:0,0.5,1,2,4,8 ng/ml
Reagent Reagent Reagent Reagent preparation preparation preparation preparation
20×wash solution:Dilute with Distilled or deionized water 1:20.
AAAAssay ssay ssay ssay procedure procedure procedure procedure
1. Prepare all r e a g e n ts before starting assay procedure. It is recommended that
all Standards and Samples be added in duplicate to the Microelisa Stripplate.
2. Add standard: Set Standard wells, testing sample wells. Add standard 50μl to
standard well.
3. Add Sample: Add testing sample 10μl then add Sample Diluent 40μl to testing
sample well; Blank well doesn’t add anyting.
4. Add 100μl of HRP-conjugate reagent to each well, cover with an adhesive strip
and incubate for 60 minutes at 37°C.
5. Aspirate each well and wash, repeating the process four times for a total of five
washes. Wash by filling each well with Wash Solution (400μl) using a squirt bottle,
manifold dispenser or autowasher. Complete removal of liquid at each step is
essential to good performance. After the last wash, remove any remaining Wash
Solution by aspirating or decanting. Invert the plate and blot it against clean paper
towels.
6. Add chromogen solution A 50μl and chromogen solution B 50μl to each well.
Gently mix and incubate for 15 minutes at 37°C. Protect Protect Protect Protect from from from from light light light light....
7. Add 50μl Stop Solution to each well. The color in the wells should change
from blue to yellow. If the color in the wells is green or the color change does not
appear uniform, gently tap the plate to ensure thorough mixing.
8. Read the Optical Density (O.D.) at 450 nm using a microtiter plate reader
within 15 minutes. CCCCalculation alculation alculation alculation of of of of results results results results
1.  This standard curve is used to determine the amount in an unknown sample.
The standard curve is generated by plotting the average O.D. (450 nm)
obtained for each of the six standard concentrations on the vertical (Y) axisversus the corresponding concentration on the horizontal (X) axis.
2. First, calculate the mean O.D. value for each standard and sample. All O.D.
values, are subtracted by the mean value of the zero standard before resul
interpretation. Construct the standard curve using graph paper or statistica
software.
3. To determine the amount in each sample, first locate the O.D. value on the
Y-axis and extend a horizontal line to the standard curve. At the point of
intersection, draw a vertical line to the X-axis and read the corresponding
concentration.
4. Any variation in operator, pipetting and washing technique, incubation time o
temperature, and kit age can cause variation in result. Each user should obtain
their own standard curve.
5. The sensitivity by this assay is 0.1 ng/ml
6. Standard curve
Storage： 2-8℃. validity： six months.
FOR FOR FOR FOR RESEARCH RESEARCH RESEARCH RESEARCH USE USE USE USE ONLY; ONLY; ONLY; ONLY; NOT NOT NOT NOT FOR FOR FOR FOR THERAPEUTIC THERAPEUTIC THERAPEUTIC THERAPEUTIC OROROROR
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