Mouse SMAD3 (Mothers against decapentaplegic homolog 3) ELISA kit

Mfr.No.

EM2177

Sensitivity

18.75pg/ml

Detection Range

31.25-2000pg/ml

Reactivity

Mouse

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 SMAD3 antibody. Add standard and properly diluted sample into relevant well respectively. After incubation, wash unbound components. Add biotinylated detection antibody. Then, it binds with SMAD3 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 tued yellow after adding a stop solution. Read the O.D. absorbance at 450nm in a microplate reader. Calculate the concentration of SMAD3 in the sample by plotting standard curve. The concentration of the target substance is proportional to the OD450 value.

Alternative Names

Mothers against decapentaplegic homolog 3 ELISA Kit, MAD homolog 3 ELISA Kit, Mad3 ELISA Kit, Mothers against DPP homolog 3 ELISA Kit, mMad3 ELISA Kit, SMAD family member 3 (SMAD 3; Smad3) ELISA Kit, Smad3 ELISA Kit, Madh3 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 SMAD3 into the sample. Calculate the recovery by comparing the measured value with the expected amount of SMAD3 in the sample.

Sample Type	Recovery Range(%)	Average(%)
serum(n=10)	88-105	98
EDTA plasma(n=10)	87-104	94
Heparin plasma(n=10)	85-103	96

Linearity

Dilute the sample with a certain amount of SMAD3 at 1:2, 1:4 and 1:8 to get the recovery range.

Sample Type	1:2	1:4	1:8
serum(n=10)	86-105%	84-96%	82-98%
EDTA plasma(n=10)	87-104%	87-99%	81-98%
Heparin plasma(n=10)	86-101%	83-88%	83-92%

Specificity

Specifically binds with SMAD3 , 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.(pg/ml)	OD-1	OD-2	Average
0	0.113	0.117	0.115
31.25	0.144	0.148	0.146
63	0.182	0.188	0.185
125	0.3	0.308	0.304
250	0.491	0.505	0.498
500	0.908	0.934	0.921
1000	1.374	1.414	1.394
2000	2.078	2.138	2.108

Sample Type

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

Assay Procedure Summary

Step 1:
Add 100ul standard or sample into each well, seal the plate and statically incubate for 90 minutes at 37°C.

Washing:
Wash the plate twice without immersing.

Step 2:
Add 100ul biotin-antibody working solution, seal the plate and statically incubate for 60 minutes at 37°C.

Washing:
Wash the plate three times and immerse for 1min each time.

Step 3:
Add 100ul HRP-Streptavidin Conjugate (SABC) working solution, seal the plate and statically incubate for 30 minutes at 37°C.

Washing:
Wash the plate five times and immerse for 1min each time.

Step 4:
Add 90ul TMB substrate solution, seal the plate and statically incubate for 10-20 minutes at 37°C. (Accurate TMB visualization control is required.)

Step 5:
Add 50ul stop solution. Read at 450nm immediately and calculate.

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 (pg/ml)	59.97	249.4	978.7	63.28	258.1	982.5
Standard deviation	2.4	10.55	42.08	3.39	10.14	39.1
CV(%)	4.01	4.23	4.3	5.35	3.93	3.98

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

Specifically binds with SMAD3 , no obvious cross reaction with other analogues.

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	Q8BUN5

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

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SKU	Supplier No.	Size	Your price	Quantity
BHE10810096	EM2177	48T	$280		Add to Cart
BHE10810096	EM2177	96T	$400		Add to Cart

Mouse SMAD3 (Mothers against decapentaplegic homolog 3) ELISA kit

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