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FOR RESEARCH USE ONLY
Drug Names
Generic Name：人中性粒细胞明胶酶相关脂质运载蛋白（NGAL）检测试剂盒
This kit can be used for determination of serum, plasma and liquid samples Organization Content.
The experimental principle:
The product levels were measured in samples of the kit by double antibody sandwichmethod. The product with the purified antibody coated microtiter plate, made of solid phase antibody, to package is the product antigen monoclonal antibodies are then added to the micropores, the product and then with HRP labeled antibody binding, the formation of antibody - antigen - antibody complex enzyme label, after thorough washing with TMB chromogenic substrate. TMB in the HRP enzyme catalytic conversion into the blue, and in the action of acid into the final yellow. This product is positively related to the depth of color and in the samples. Instrument measured absorbance in the 450nm wavelength with ELISA (OD), the product concentration in the samples was calculated by standard curve.
Materials provided with the kit
Materials provided with the kit 48determinations 96 determinations Storage
User manual 1 1
Closure plate membrane 2 2
Sealed bags 1 1
Microelisa stripplate 1 1 2-8℃
Standard：360ng/L 0.5ml×1 bottle 0.5ml×1 bottle 2-8℃
Standard diluent 1.5ml×1 bottle 1.5ml×1 bottle 2-8℃
HRP-Conjugate reagent 3ml×1 bottle 6ml×1 bottle 2-8℃
Sample diluent 3ml×1 bottle 6ml×1 bottle 2-8℃
Chromogen Solution A 3ml×1 bottle 6ml×1 bottle 2-8℃
Chromogen Solution B 3ml×1 bottle 6ml×1 bottle 2-8℃
Stop Solution 3ml×1 bottle 6ml×1 bottle 2-8℃
wash solution （20ml×20 fold）
×1bottle （20ml×30 fold）
×1bottle 2-8℃
人中性粒细胞明胶酶相关脂质运载蛋白（NGAL）检测试剂盒Specimen requirements
1.serum- coagulation at room temperature 10-20 mins，centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
2.plasma-use suited EDTA or citrate plasma as an anticoagulant,mix 10-20 mins ,centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
3.Urine-collect sue a sterile container, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again. The Operation of Hydrothorax and cerebrospinal fluid Reference to it.
4.cell culture supernatant-detect secretory components, collect sue a sterile container, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant,detect the composition of cells, Dilut cell suspension with PBS（PH7.2-7.4）, Cell concentration reached 1 million / ml, repeated freeze-thaw cycles, damage cells and release of intracellular components, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
5.Tissue samples- After cutting samples, check the weight,add PBS（PH7.2-7.4）, Rapidly frozen with liquid nitrogen, maintain samples at 2-8℃ after melting,add PBS（PH7.4）, Homogenized by hand or Grinders, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant.
6.extract as soon as possible after Specimen collection,and according to the relevant literature, and should be experiment as soon as possible after the extraction. If it can’t, specimen can be kept in -20 ℃ to preserve, Avoid repeated freeze-thaw cycles.
7.Can’t detect the sample which contain NaN3, because NaN3 inhibits HRP active.
Assay procedure
1.Dilute and add sample to Standard: set 10 Standard wells on the ELISA plates coated, add Standard 100μl to the first and the second well, then add Standard dilution 50μl to the first and the second well, mix; take out 100μl form the first and the second well then add it to the third and the forth well separately. then add Standard dilution 50μl to the third and the forth well ,mix ; then take out 50μl from the third and the forth well discard, add 50μl to the fifth and the sixth well ,then add Standard dilution 50μl to the fifth and the sixth well, mix ; take out 50μl from the fifth and the sixth well and add to the seventh and the eighth well, then add Standard dilution 50μl to the seventh and the eighth well ,mix ; take out 50μl from the seventh and the eighth well and add to the ninth and the tenth well, add Standard dilution 50μl to the ninth and the tenth well, mix , take out 50μl from the ninth and the tenth well discard(add Sample 50μl to each well after Diluting ,(density: 240ng/L,160ng/L ,80ng/L,40ng/L, 20ng/L)
2.add sample：Set blank wells separately (blank comparison wells .don’t add sample and HRP-Conjugate reagent, other each step operation is same). testing sample well. add Sample dilution 40μl to testing sample well, then add testing sample 10μl (sample final dilution is 5-fold), add sample to wells , don’t touch the well wall as far as possible, and Gently mix.
3.Incubate: After closing plate with Closure plate membrane ,incubate for 30 min at 37℃.
4.Configurate liquid: 30-fold (or 20-fold)wash solution diluted 30-fold (or 20-fold) with distilled water and reserve.
5.washing：Uncover Closure plate membrane, discard Liquid, dry by swing, add washing buffer to every well, still for 30s then drain, repeat 5 times, dry by pat.
6.add enzyme：Add HRP-Conjugate reagent 50μl to each well, except blank well. 
7.incubate：Operation with 3.
8.washing：Operation with 5.
9.color：Add Chromogen Solution A 50ul and Chromogen Solution B to each well, evade the light preservation for 15 min at 37℃
10.Stop the reaction：Add Stop Solution50μl to each well, Stop the reaction(the blue color change to yellow color).
11.assay：take blank well as zero , Read absorbance at 450nm after Adding Stop Solution and within 15min.
Important notes
1.The kit takes out from the refrigeration environment should be balanced 15-30 minutes in the room temperature, ELISA plates coated if has not use up after opened, the plate should be stored in Sealed bag.
2.washing buffer will Crystallization separation, it can be heated the water helps dissolve when dilute . Washing does not affect the result.
3.add Sample with sampler Each step, And proofread its accuracy frequently, avoids the experimental error. add sample within 5 mins, if the number of sample is much , recommend to use Volley .
4.if the testing material content is excessively higher (The sample OD is bigger than the first standard well ),please dilute Sample (n-fold), Please diluente and multiplied by the dilution factor.（×n×5）.
5.Closure plate membrane only limits the disposable use, to avoid cross-contamination.
6.The substrate evade the light preservation.
7.Please according to use instruction strictly, The test result determination must take the microtiter plate reader as a standard.
8.All samples, washing buffer and each kind of reject should according to infective material process.
9.Do not mix reagents with those from other lots.
Calculate：
Take the standard density as the horizontal, the OD value for the vertical ,draw the standard curve on graph paper, Find out the corresponding density according to the sample OD value by the Sample curve, multiplied by the dilution multiple, or calculate the straight line regression equation of the standard curve with the standard density and the OD value ,with the sample OD value in the equation, calculate the sample density, multiplied by the dilution factor, the result is the sample actual density.
Storage and validity
1．Storage： 2-8℃.
2．validity： six months.
人中性粒细胞明胶酶相关脂质运载蛋白（NGAL）检测试剂盒其它产品
TF4 酸 100 mg
TF4 胺 5 mg
TF4 马来酰亚胺 5 mg
TF4 亚磷酰胺 1 g
TF4 琥珀酰亚胺酯 25 mg
台盼兰 5 g
台盼兰 25 g
台盼兰 100 g
台盼红（0.1 M水溶液） 1 mL
台盼红（0.1 M水溶液） 10 mL
酚红钠盐 5 g
酚红钠盐 25 g
B-藻红蛋白 1 mg
交联藻蓝蛋白 1 mg
C-藻蓝蛋白 1 mg
别藻蓝蛋白 1 mg
R-藻红蛋白 1 mg
6-氨基己醇 25 g
6-氨基己醇 100 g
N-BOC-尸胺 5 g
N-BOC-尸胺 25 g
N-BOC-乙二胺 5 g
N-BOC-乙二胺 25 g
N-FMOC-乙二胺 5 g
N-FMOC-乙二胺 25 g
N-FMOC-尸胺 5 g
N-FMOC-尸胺 25 g
溴甲基乙酸盐 5 g
3,6-二氯偏苯三酸酐 1 g
MMT保护-5'-氨基改良剂C6 1 g
TFA保护-5'-氨基改良剂C6 1 g
4-(N-马来酰亚胺基甲基)环己烷-1-羧酸琥珀酰亚胺酯 5 mg
4-(N-马来酰亚胺基甲基)环己烷-1-羧酸琥珀酰亚胺酯 25 mg
4-(N-马来酰亚胺基甲基)环己烷-1-羧酸琥珀酰亚胺酯 100 mg
4-(N-马来酰亚胺基甲基)环己烷-1-羧酸-3-磺基琥珀酰亚胺酯 10 mg
4-(N-马来酰亚胺基甲基)环己烷-1-羧酸-3-磺基琥珀酰亚胺酯 50 mg
 3-马来酰亚胺丙酸琥珀酰亚胺酯 100 mg
FMOC-Asp(EDANS)-OH 1 g
FMOC-Asp(EDANS)-OH 5 g
FMOC-Asp(5/6-FAM)-OH 1 g
FMOC-Asp(5-FAM)-OH 1 g
FMOC-Asp(5/6-TAMRA)-OH 1 g
FMOC-Asp(5-TAMRA)-OH 1 g
FMOC-Glu(EDANS)-OH 1 g
FMOC-Glu(EDANS)-OH 5 g
FMOC-Glu(5/6-FAM)-OH 1 g
FMOC-Glu(5-FAM)-OH 1 g
FMOC-Glu(5/6-TAMRA)-OH 1 g
FMOC-Glu(5-TAMRA)-OH 1 g
FMOC-Lys(DABCYL)-OH 1 g
FMOC-Lys(DABCYL)-OH 5 g
FMOC-Lys(5/6-FAM)-OH 1 g
FMOC-Lys(5-FAM)-OH 1 g
FMOC-Lys(5/6-TAMRA)-OH 1 g
FMOC-Lys(5-TAMRA)-OH 1 g
FMOC-Lys(Tide Fluor™ 2)-OH 1 g
FMOC-Lys(Tide Quencher™ 2)-OH 1 g
BOC-Statine [Boc-Sta(3S,4S)-OH] 1 g
BOC-Statine [Boc-Sta(3S,4S)-OH] 10 g
FMOC-Statine [Boc-Sta(3S,4S)-OH] 1 g
FMOC-Statine [Boc-Sta(3S,4S)-OH] 10 g
Statine 100 mg
Stain-All 100 mg
10-乙酰基-3,7-二羟基吩嗪 25 mg
ReadiUse™ 过氧化氢片剂 25 Ct
ReadiUse™ 过氧化氢溶液 5x10 mL
Amplite™蓝 5 mg
Amplite™ IR 1 mg
HRP 交联稳定剂 50 mL
鲁米诺；3-氨基苯二甲酰肼；发光氨 5 g
鲁米诺；3-氨基苯二甲酰肼；发光氨 25 g
荧光素二磷酸酯四铵盐 5 mg
4-甲基伞形酮磷酸酯二钠盐 100 mg
4-甲基伞形酮磷酸酯二钠盐 5 g
4-甲基伞形酮磷酸酯 100 mg
4-甲基伞形酮磷酸酯 5 g
Ac-DEVD-AFC 5 mg
Ac-DEVD-AMC 5 mg
Ac-DEVD-CHO 5 mg
Ac-DEVD-pNA 5 mg
FITC-C6-DEVD-FMK 1 mg
FITC-C6-LEHD-FMK 1 mg
Ac-IETD-AFC 5 mg
Ac-IETD-AMC 5 mg
Ac-IETD-CHO 5 mg
Z-IETD-pNA 5 mg
D-氨基虫荧光素 10 mg
Z-DEVD-AFC 5 mg
Z-DEVD-AMC 5 mg
Z-DEVD-pNA 5 mg
Z-IETD-AFC 5 mg
(Z-DEVD)2-R110 5 mg
(Z-IETD)2-R110 5 mg
FITC-酪蛋白交联试剂 25 mg
TAMRA-酪蛋白交联试剂 5 mg
Gly-Pro-AMC 10 mg
cAMP-SP-TAMRA （红色荧光PDE IV底物） 1 mg
cGMP-SP-TAMRA （红色荧光PDE V底物） 1 mg
FDG 5 mg
FDGlcU 5 mg
CUG 10 mg
MUG 1 g
MUG 10 g
4-甲基伞形酮-β-D-葡糖苷 250 mg
MUGlcU 100 mg
7-羟基吩恶嗪酮 β-D-吡喃型半乳糖 25 mg
3-氰基-7-乙氧基香豆素 10 mg
7-乙氧基-4-三氟甲基香豆素 25 mg
7-苄氧基试卤灵 10 mg
7-羟基吩恶嗪酮; 乙氧基试卤灵 5 mg
甲氧基试卤灵 5 mg
7-羟基吩恶嗪酮二戊醚 5 mg
二氢乙锭 25 mg
二氢乙锭（5 mM DMSO溶液） 1 mL
二氢荧光素二乙酸盐 500 mg
2',7'-二氯荧光素二乙酸酯 100 mg
MCLA 5 mg
刃天青钠盐 1 g
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