Rat HDGF (Hepatoma Derived Growth Factor) CLIA Kit (RTES00294)

SKU:
RTES00294
€649

Description

ELISA Kit Technical ManualMSDS

Rat HDGF (Hepatoma Derived Growth Factor) CLIA Kit

This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Rat HDGF . Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Rat HDGF and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Rat HDGF, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Rat HDGF. The concentration of Rat HDGF in the samples can be calculated by comparing the RLU of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityRat
Detection methodChemiluminescence
Detection range62.50-4000 pg/mL
Sensitivity37.50 pg/mL
Sample volume100uL
Sample typeSerum, plasma and other biological fluids
RepeatabilityCV < 15%

Specificity

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

Typical Data

As the RLU 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)
RLU Average Corrected
400055253
62909
5908159046
200026356
29362
2785927824
100014664
12304
1348413449
5006320
6890
66056570
2503330
3156
32433208
1251598
1564
15811546
62.50736
774
755720
034
36
35--

Precision

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

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

  Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)217.85637.881675.71219.88649.981576.67
Standard deviation20.3564.81179.6420.1676.24154.20
C V (%)9.3410.1610.729.1711.739.78

Recovery

The recovery of Rat HDGF 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)92-10799
EDTA plasma (n=5)91-10598
Cell culture media (n=5)98-111105

Linearity

Samples were spiked with high concentrations of Rat HDGF 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 (%)94-10789-103100-116
Average (%)10094106
1:4Range (%)101-119101-11890-101
Average (%)11010895
1:8Range (%)86-101103-11898-109
Average (%)93110104
1:16Range (%)87-9886-9789-100
Average (%)939294

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 CLIA Plate(Dismountable)8 wells X12 strips-20'C, 6 months
Reference Standard2 vials
Concentrated Biotinylated Detection Ab (100X)1 vial, 120 uL
Concentrated HRP Conjugate (100X)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 (25X)1 vial, 30 mL
Substrate Reagent A1 vial, 5 mL4'C (shading light)
Substrate Reagent B1 vial, 5 mL4'C (shading light)
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Rat HDGF (Hepatoma Derived Growth Factor) CLIA Kit (RTES00294) Assay procedure

    1. Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
    2. Aliquot 100µl of standard solutions into the standard wells.
    3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids.) into test sample wells.
    5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
    6. Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix.Incubate for 1 hour at 37°C.
    7. Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
    8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
    9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
    10. Add 100µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37°C. Protect the plate from light.
    11. Determine the RLU value of each well immediately.

Rat HDGF (Hepatoma Derived Growth Factor) CLIA Kit (RTES00294) Protein Information

UniProt Protein Function:HDGF: Heparin-binding protein, with mitogenic activity for fibroblasts. Acts as a transcriptional repressor. Belongs to the HDGF family.
UniProt Protein Details:

Protein type:Cytokine; Transcription regulation

Cellular Component: nucleoplasm; extracellular space; transcriptional repressor complex; cytoplasm; nucleus

Molecular Function:heparin binding; growth factor activity; nucleotide binding

Biological Process: transcription, DNA-dependent; positive regulation of transcription from RNA polymerase II promoter; negative regulation of transcription from RNA polymerase II promoter; cellular process

NCBI Summary:induces DNA replication in vascular smooth muscle cells [RGD, Feb 2006]
UniProt Code:Q8VHK7
NCBI GenInfo Identifier:16758528
NCBI Gene ID:114499
NCBI Accession:NP_446159.1
UniProt Secondary Accession:Q8VHK7,Q923W3,
UniProt Related Accession:Q8VHK7
Molecular Weight:26,488 Da
NCBI Full Name:hepatoma-derived growth factor
NCBI Synonym Full Names:hepatoma-derived growth factor
NCBI Official Symbol:Hdgf  
NCBI Protein Information:hepatoma-derived growth factor
UniProt Protein Name:Hepatoma-derived growth factor
Protein Family:Hepatoma-derived growth factor
UniProt Gene Name:Hdgf  
UniProt Entry Name:HDGF_RAT
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Additional Information

Product type:
ELISA
Reactivity:
Rat
ELISA Type:
CLIA
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