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BH4(Tetrahydrobiopterin) ELISA Kit (UNES00059)

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SKU:
UNES00059
$567

Description

BH4(Tetrahydrobiopterin) ELISA Kit

This ELISA kit uses Competitive-ELISA as the method. The microtiter plate provided in this kit has been pre-coated with BH4. During the reaction, BH4 in the sample or standard competes with a fixed amount of  BH4 on the solid phase supporter for sites on the Biotinylated Detection Ab specific to  BH4. Excess conjugate and unbound sample or standard are washed from the plate, and Avidin conjugated to Horseradish Peroxidase (HRP) are added to each microplate well and incubated. Then a TMB substrate solution is added to each well. The enzyme-substrate reaction is terminated by adding Stop Solution and the color change is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The concentration of   BH4 in the samples is then determined by comparing the OD of the samples to the standard curve.

Assay typeCompetitive-ELISA
Format96T
Assay time2.5h
ReactivityUniversal
Detection MethodColormetric
Detection Range31.25-2000 pg/mL
Sensitivity18.75 pg/mL
Sample Volume50uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes BH4 in samples. No significant cross-reactivity or interference between BH4 and analogues was observed.

Typical Data

As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.

Concentration(pg/mL) O.D Average
20000.274
0.322
0.298
10000.359
0.409
0.384
5000.55
0.542
0.546
2500.812
0.826
0.819
1251.204
1.198
1.201
62.51.622
1.62
1.621
31.251.967
1.981
1.974
02.477
2.489
2.483

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level BH4 were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level BH4 were tested on 3 different plates, 20 replicates in each plate.

  Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)105.95238.37995.86112.62233.331067.77
Standard deviation6.7913.4050.396.2713.9150.83
C V (%)6.415.625.065.575.964.76

Recovery

The recovery of BH4 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type Range (%) Average Recovery (%)
Serum (n=5)89-10496
EDTA plasma (n=5)88-10396
Cell culture media (n=5)93-108100

Linearity

Samples were spiked with high concentrations of BH4 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.

    Serum (n=5) EDTA plasma (n=5) Cell culture media (n=5)
1:2Range (%)93-10793-10891-106
Average (%)1009997
1:4Range (%)89-10394-10988-101
Average (%)9610093
1:8Range (%)94-10788-10090-103
Average (%)1019498
1:16Range (%)89-10194-10687-97
Average (%)9610092

Kit Components & Storage

An unopened kit can be stored at 4'C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.

Item Specifications Storage
Micro ELISA Plate(Dismountable)8 wells ×12 strips-20'C, 6 months
Reference Standard2 vials
Concentrated Biotinylated Detection Ab (100×)1 vial, 120 uL
Concentrated HRP Conjugate (100×)1 vial, 120 uL-20'C(shading light), 6 months
Reference Standard & Sample Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25×)1 vial, 30 mL
Substrate Reagent1 vial, 10 mL4'C(shading light)
Stop Solution1 vial, 10 mL4'C
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy
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Additional Information

Reactivity:
Universal
ELISA Type:
Competitive
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