Rat Hspa5 (78 kDa glucose-regulated protein) ELISA Kit

Mfr.No.

ER0562

Sensitivity

0.188ng/ml

Detection Range

0.313-20ng/ml

Reactivity

Rat

Assay Duration

4 hours

Assay Type

Sandwich

Description

This kit is based on Double antibody-Sandwich ELISA detection method and takes 4h assay time. The microplate provided in this kit has been precoated with anti Hspa5 antibody. Add standard and properly diluted sample into relevant well respectively. After incubation, wash unbound components. Add biotinylated detection antibody. Then, it binds with Hspa5 bound to precoated antibody. Wash unbound components and add HRP-Streptavidin Conjugate (SABC). Wash unbound components again and add TMB substrate solution. Then, TMB was catalyzed by HRP to produce a blue color product that turned yellow after adding a stop solution. Read the O.D. absorbance at 450nm in a microplate reader. Calculate the concentration of Hspa5 in the sample by plotting standard curve. The concentration of the target substance is proportional to the OD450 value.

Alternative Names

Endoplasmic reticulum chaperone BiP ELISA Kit, 78 kDa glucose-regulated protein ELISA Kit, GRP-78 ELISA Kit, Binding-immunoglobulin protein(BiP ELISA Kit, Heat shock protein 70 family protein 5(HSP70 family protein 5 ELISA Kit, Heat shock protein family A member 5 ELISA Kit, Immunoglobulin heavy chain-binding protein ELISA Kit, Steroidogenesis-activator polypeptide ELISA Kit, HSPA5 ELISA Kit, GRP78 ELISA Kit

Detection Method

Double Antibody

Kit Components

Kit Components	Item	Size(48T)	Size(96T)	Storage Condition for Opened Kit
E001	ELISA Microplate(Dismountable)	8x6	8x12	Put the rest strips into a sealed foil bag with the desiccant. Stored for 1 month at 2-8°C; Stored for 6 month at -20°C
E002	Lyophilized Standard	1vial	2vial	Put the rest standards into a desiccant bag. Stored for 1 month at 2-8°C; Stored for 6 month at -20°C
E003	Biotin-labeled Antibody(Concentrated, 100X)	60ul	120ul	2-8°C (Avoid Direct Light)
E034	HRP-Streptavidin Conjugate(SABC, 100X)	60ul	120ul
E024	TMB Substrate	5ml	10ml
E039	Sample Dilution Buffer	10ml	20ml	2-8°C
E040	Antibody Dilution Buffer	5ml	10ml
E049	SABC Dilution Buffer	5ml	10ml
E026	Stop Solution	5ml	10ml
E038	Wash Buffer(Concentrated, 25X)	15ml	30ml
E006	Plate Sealer	3 pieces	5 pieces
E007	Product Description	1 copy	1 copy

Recovery

Add a certain amount of Hspa5 into the sample. Calculate the recovery by comparing the measured value with the expected amount of Hspa5 in the sample.

Sample Type	Recovery Range(%)	Average(%)
serum(n=10)	87-99	95
EDTA plasma(n=10)	85-105	97
Heparin plasma(n=10)	86-99	90

Linearity

Dilute the sample with a certain amount of Hspa5 at 1:2, 1:4 and 1:8 to get the recovery range.

Sample Type	1:2	1:4	1:8
serum(n=10)	92-105%	83-98%	89-97%
EDTA plasma(n=10)	87-104%	84-99%	84-95%
Heparin plasma(n=10)	87-103%	83-100%	80-91%

Specificity

Specifically binds with Hspa5 , no obvious cross reaction with other analogues.

Standard Curve

This product is detected by QC department and meets performance required in the manual. (Laboratory Humidity: 20%-60%; Temperature: 18°C -25°C; Equilibrate TMB substrate to 37°C before staining. After adding into the ELISA wells, incubate for 15min at 37°C in dark.) Due to different assay environments and operations, assay data below and standard curve are provided for reference. Experimenters should establish standard curve according to their own assay.

STD.(ng/ml)	OD-1	OD-2	Average
0	0.048	0.05	0.049
0.312	0.119	0.123	0.121
0.625	0.234	0.24	0.237
1.25	0.414	0.426	0.42
2.5	0.634	0.652	0.643
5	1.109	1.141	1.125
10	1.646	1.694	1.67
20	2.252	2.318	2.285

Sample Type

Tissue lysate, Cell lysates, Cell culture supernatants, Plasma, Other biological fluids

Components

Microplate,Lyophilized Standard,buffer,SABC,Label antibody,SABC,TMB,stop solution

Precision

Intra-assay Precision: samples with low, medium and high concentration are tested 20 times on the same plate.
Inter-assay Precision: samples with low, medium and high concentration are tested 20 times on three different plates.

Item	Intra-assay Precision			Inter-assay Precision
Sample	1	2	3	1	2	3
n	20	20	20	20	20	20
Mean (ng/ml)	0.64	2.52	10.19	0.65	2.46	10.17
Standard deviation	0.04	0.16	0.52	0.03	0.12	0.53
CV(%)	5.82	6.42	5.12	4.89	4.69	5.23

Background

Hspa5/GRP-78/Grp78/Steroidogenesis-activator polypeptide/Heat shock 70 kDa protein 5/Immunoglobulin heavy chain-binding protein(BiP)

Stability

Perform the stability test for the sealed kit at 37°C and 2-8°C and get relevant data.

ELISA kit(n=5)	37°C for 1 month	2-8°C for 6 months
Average(%)	80	95-100

Notes

For Research Use Only

Usage Notes

Step 1:
Take out the required plate wells, add 50ul Cap/Det Ab into each well, then add 50ul Standard or Sample into individual well. (When adding standard or sample, the disposable tip lightly touches the liquid level. Change the disposable tips for different samples and standards.) Gently tap the plate for 10s to ensure thorough mixing then static incubate for 60 minutes at 37°C.

Washing:
Wash the plate twice without immersion.

Step 2:
Add 100ul HRP-Streptavidin (orange) into each well, seal the plate and static incubate for 30 minutes at 37°C.

Washing:
Wash the plate five times without immersion.

Step 3:
Add 90ul TMB substrate solution, seal the plate and static incubate for 10-20 minutes at 37°C. (Accurate TMB visualization control is required.)

Step 4:
Add 50ul stop solution. Read at 450nm immediately and calculate.

UniProt ID	P06761

Applications	ELISA
Applications Notes	Hspa5 ELISA Kit allows for the in vitro quantitative determination of Hspa5 concentrations in serum, plasma, tissue homogenates and other biological fluids.

Storage	2-8°C(Sealed), Don't cryopreserve.

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Rat Hspa5 (78 kDa glucose-regulated protein) ELISA Kit

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