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Human NSE 酶联免疫分析（ELISA）
试剂盒使用说明书
货号: HY10786
本试剂盒用于体外定量检测人血清、血浆、组织、细胞上清及相关液体样本中NSE的含量。
本试剂盒用于体外定量检测人血清、血浆、组织、细胞上清及相关液体样本中NSE的含量。
本试剂盒应用双抗体夹心法测定标本中人NES水平。用纯化的人NES抗体包被微孔板，制成固相抗体，往包被单抗的微孔中依次加入NES，再与HRP标记的NSE抗体结合，形成抗体-抗原-酶标抗体复合物，经过*洗涤后加底物TMB显色。TMB在HRP酶的催化下转化成蓝色，并在酸的作用下转化成zui终的黄色。颜色的深浅和样品中的NSE呈正相关。用酶标仪在450nm波长下测定吸光度（OD值），通过标准曲线计算样品中人NSE浓度。
l 本试剂盒仅用于科研，不得用于医学诊断。
l 使用试剂盒前，请仔细阅读说明书，以试剂盒内的说明书为准，请务必理解说明书内容。
l 酶标包被板属于可拆型，开封后如未用完，板条应装入铝箔袋密封并2°C-8°C保存。
l 加样样本和试剂应尽快，每一步加样都根据相同的序列。Pipetting of samples and reagents must be done as quickly as possible and in the same sequence for each step.
l Use reservoirs only for single reagents. This especially applies to the substrate reservoirs. Using a reservoir for dispensing a substrate solution that had previously been used for the conjugate solution may turn solution colored. Do not pour reagents back into vials as reagent contamination may occur.
l Mix the contents of the microplate wells thoroughly to ensure good test results. Do not reuse microwells.
l 避免实验过程中酶标板干燥；清洗后尽快加试剂。Do not let wells dry during assay; add reagents immediay after completing the rinsing steps.
l 实验开始，所有试剂应平衡到室温 （21-26°C） 后方可进行。温度会影响吸光度值，但不影响样本值。Allow the reagents to reach room temperature （21-26°C） before starting the test. Temperature will affect the absorbance readings of the assay. However, values for the patient samples will not be affected.
l 切忌用口加样，以免试剂和样本粘到皮肤和粘膜上。Never pipet by mouth and avoid contact of reagents and specimens with skin and mucous membranes.
l 不要在处理样本或试剂的区域抽烟、吃东西、喝酒或化妆。Do not smoke, eat, drink or apply cosmetics in areas where specimens or kit reagents are handled.
l 操作时带一次性手套，微生物的污染会影响实验结果的正确性。Wear disposable latex gloves when handling specimens and reagents. Microbial contamination of reagents or specimens may give false results.
l 操作应按照国家规定的微生物安全条列进行。Handling should be done in accordance with the procedures defined by an appropriate national biohazard safety guideline or regulation.
l 过期的试剂盒切勿再使用。Do not use reagents beyond expiry date as shown on the kit labels.
l All indicated volumes have to be performed according to the protocol. Optimal test results are only obtained when using calibrated pipettes and microtiterplate readers.
l Do not mix or use components from kits with different lot numbers. It is advised not to exchange wells of different plates even of the same lot. The kits may have been shipped or stored under different conditions and the binding characteristics of the plates may result slightly different.
l Avoid contact with Stop Solution containing 0.5 M H2SO4. It may cause skin irritation and burns.
l 含有防腐剂、BND或者类似防腐剂MIT的试剂Some reagents contain Proclin, BND and/or MIT as preservatives. In case of contact with eyes or skin, flush immediay with water.
l TMB 底物对皮肤有刺激性，不慎入眼，立即用大量水冲洗，皮肤上不慎接触到也应立即用肥皂，并用大量的水冲洗。试验中被污染的物品在下次使用前应尽快清洗。TMB substrate has an irritant effect on skin and mucosa. In case of possible contact, wash eyes with an abundant volume of water and skin with soap and abundant water. Wash contaminated objects before reusing them. If inhaled, take the person to open air.
l 配置的化学品或使用后的试剂，根据国家微生物安全条列都视为危险废物。Chemicals and prepared or used reagents have to be treated as hazardous waste according to the national biohazard safety guideline or regulation.
l 参照材料安全系数表知道试剂盒里的有害物质。For information on hazardous substances included in the kit please refer to Material Safety Data Sheets
| 说明书 | 1 | |
| | 2 | |
| 酶标包被板 | 12孔×8条 | 2-8℃ |
| 标准 | 0.5ml×1 瓶 0 ng/ml 0.5ml×1瓶 6.0 ng/ml 0.5ml×1瓶 12 ng/ml 0.5ml×1瓶 24 ng/ml 0.5ml×1瓶 48 ng/ml 0.5ml×1瓶 96 ng/m | 2-8℃ |
| 样本亲和素 | 1.5ml×1瓶 | 2-8℃ |
| 显色剂A液 | 6ml×1瓶 | 2-8℃ |
| 显色剂B液 | 6ml×1瓶 | 2-8℃ |
| 终止液 | 6ml×1瓶 | 2-8℃ |
| 酶标试剂 | 6ml×1瓶 | 2-8℃ |
| 30倍浓缩洗涤液 | 20ml×1瓶 | 2-8℃ |
l 吸光度值在450nm波长下检测的酶标仪。
l 1-2ml的加样器。
l 100 ml和1L的量筒。
l Calibrated adjustable precision pipettes, preferably with disposable plastic tips. （A manifold multi-channel pipette is desirable for large assays.）
l 吸水纸。
l 37°C 恒温箱。
l 蒸馏水或去离子水。
l 数据分析和绘图软件，图纸：线性（笛卡尔），Data analysis and graphing software. Graph paper: linear （Cartesian）,log-log or semi-log, or log-logit as desired.
l 标准和样品稀释的试管。Tubes to prepare standard or sample dilutions.
u 有效期内的试剂如开封后未用完，请2-8°C 保存。
u 过期的试剂请不要用，打开后的试剂2-8°C 保存。
u 酶标包被板必须2-8°C 保存，如有未用完的酶标包被板，打开的铝箔袋须密封并2-8°C 保存。
u 开封后的试剂盒2-8°C只能保存8周。
试剂盒从冷藏环境中取出应在室温平衡后方可使用。
血清-用采血管收集血液，室温血液自然凝固30分钟，离心15分钟左右（1000转/分），收集上清，尽早进行实验，或者分装后-20°C 或 -80°C 保存。Use a serum separator tube（SST） and allow samples to clot for 30minutes before centrifugation for 15minutes at approximay 1000 xg.Remove serum and assay immediay or aliquot and store samples at -20°C or -80°C.
血浆-收集好的血浆根据要求选择EDTA或者肝素作为抗凝剂，2-8°C离心15分钟左右（1000转/分），30分钟内收集好，-20°C 或 -80°C 保存，避免反复冻融。
细胞上清及相关液体-收集好的样本离心后应尽快实验，或者分装后-20°C 或 -80°C 保存，避免反复冻融。
u 注意事项
l 所有试剂和样本使用前必须在室温下平衡，混匀试剂时不应起泡沫。All reagents and specimens must be allowed to come to room temperature before use. All reagents must be mixed without foaming.
l 一旦实验开始，各操作步骤都应尽快完成。Once the test has been started, all steps should be completed without interruption.
l 避免重复使用手中的吸头和试管，以免交叉污染。Use new disposal plastic pipette tips for each standard, control or sample in order to avoid cross contamination.
l 吸光度值是温育时间和温度的函数，实验开始前，准备好所有的试剂，Absorbance is a function of the incubation time and temperature. Before starting the assay, it is recommended that all reagents are ready, caps removed, all needed wells secured in holder, etc. This will ensure equal elapsed time for each pipetting step without interruption.
l 一般情况，酶的反应与时间和温度成正比。As a general rule the enzymatic reaction is linearly proportional to time and temperature.
l Determine absorption with an ELISA reader at 450 nm against 620 nm as reference. If no reference wavelength is available, read only at 450nm. If the extinction of the highest standard exceeds the measurement range of the photometer, absorption must be measured immediay at 405 nm against 620 nm as reference.
u 操作步骤
1. 加标准品：在酶标包被板上设标准品孔，依次加入不同浓度的标准品50ul（建议每个浓度做2个平行孔）。
2. 加样：分别设空白孔（空白对照孔不加样品及酶标试剂，其余各步操作相同）、待测样品孔。在酶标包被板上待测样品孔中先加样品40μl，然后再加样品亲和素10μl。加样将样品加于酶标板孔底部，尽量不触及孔壁，轻轻晃动混匀。
3. 温育：用封板膜封板后置37℃温育30分钟。
4. 配液：将30倍浓缩洗涤液用蒸馏水30倍稀释后备用。
5. 洗涤：小心揭掉封板膜，弃去液体，甩干，每孔加满洗涤液，静置30秒后弃去，如此重复5次，拍干。
6. 加酶：每孔加入酶标试剂50μl，空白孔除外。
7. 温育：操作同3。
8. 洗涤：操作同5。
9. 显色：每孔先加入显色剂A50μl，再加入显色剂B50μl，轻轻震荡混匀，37℃避光显色15分钟.
10. 终止：每孔加终止液50μl，终止反应（此时蓝色立转黄色）。
11. 测定：以空白空调零，450nm波长依序测量各孔的吸光度（OD值）。 测定应在加终止液后15分钟以内进行
l Calculate the average absorbance values for each set of standards, controls and patient samples.
l Construct a standard curve by plotting the mean absorbance obtained from each standard against its.
l Concentration with absorbance value on the vertical（Y） axis and concentration on the horizontal （X） axis.
l Using the mean absorbance value for each sample determine the corresponding concentration from the standard curve.
l Automated method: The results in the IFU have been calculated automatically using a 4 PL.
l （4 Parameter Logistics） curve fit. 4 Parameter Logistics is the preferred calculation method. Other data.
l Reduction functions may give slightly different results.
l The concentration of the samples can be read directly from this standard curve. Samples with.
l Concentrations higher than that of the highest standard have to be further diluted. For the calculation of.
l The concentrations this dilution factor has to be taken into account.
REF : Cat.-No.: / Kat.-Nr.: / No.- Cat.: / Cat.-No.: / N.º Cat.: / N.–Cat
LOT : Lot-No.: / Chargen-Bez.: / No. Lot: / Lot-No.: / Lote N.º: / Lotto n.:
:No. of Tests: / Kitgröße: / Nb. de Tests: / No. de Determ.: / N.º de Testes: / Quantità dei tests:
: Keep away from heat or direct sun light. / Vor Hitze und direkter Sonneneinstrahlung schützen. /
Garder à l’abri de la chaleur et de toute exposition lumineuse. / Manténgase alejado del calor o la
luz solar directa. / Manter longe do calor ou luz solar directa. / Non esporre ai raggi solari.
: Read instructions before use. / Arbeitsanleitung lesen. / Lire la fiche technique avant emploi. /Lea las instrucciones antes de usar. / Ler as instruções antes de usar. / Leggere le istruzioni prima dell’uso.
: Store at: / Lagern bei: / Stocker à: / Almacene a :/ Armazenar a :/ Conservare a:
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