Rat NEF3 (Neurofilament 3) ELISA Kit

Mfr.No.

ER1183

Sensitivity

18.75pg/ml

Detection Range

31.25-2000pg/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 NEF3 antibody. Add standard and properly diluted sample into relevant well respectively. After incubation, wash unbound components. Add biotinylated detection antibody. Then, it binds with NEF3 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 NEF3 in the sample by plotting standard curve. The concentration of the target substance is proportional to the OD450 value.

Alternative Names

Neurofilament medium polypeptide ELISA Kit, NF-M ELISA Kit, 160 kDa neurofilament protein ELISA Kit, Neurofilament 3 ELISA Kit, Neurofilament triplet M protein ELISA Kit, NEFM ELISA Kit, NEF3 ELISA Kit, NFM 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 NEF3 into the sample. Calculate the recovery by comparing the measured value with the expected amount of NEF3 in the sample.

Sample Type	Recovery Range(%)	Average(%)
serum(n=10)	92-105	97
EDTA plasma(n=10)	86-99	93
Heparin plasma(n=10)	87-105	93

Linearity

Dilute the sample with a certain amount of NEF3 at 1:2, 1:4 and 1:8 to get the recovery range.

Sample Type	1:2	1:4	1:8
serum(n=10)	85-99%	82-96%	82-95%
EDTA plasma(n=10)	83-97%	80-99%	82-95%
Heparin plasma(n=10)	82-98%	80-100%	83-97%

Specificity

Specifically binds with NEF3 , 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	0.148	0.152	0.15
31.25	0.28	0.288	0.284
62.5	0.374	0.384	0.379
125	0.578	0.594	0.586
250	0.781	0.803	0.792
500	1.053	1.083	1.068
1000	1.629	1.677	1.653
2000	2.201	2.265	2.233

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 (pg/ml)	61.23	262	985.8	64.54	251	991.3
Standard deviation	3.27	14.07	55.11	3.4	13.78	55.12
CV(%)	5.34	5.37	5.59	5.27	5.49	5.56

Background

NEF3/NEFM/NEF3neurofilament medium polypeptide/neurofilament, medium polypeptide/neurofilament, medium polypeptide 150kDa/neurofilament-3(150 kD medium)/NF-M160 kDa neurofilament protein/NFMNeurofilament triplet M protein

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	P12839

Applications	ELISA
Applications Notes	NEF3 ELISA Kit allows for the in vitro quantitative determination of NEF3 concentrations in serum, plasma, tissue homogenates and other biological fluids.

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

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Rat NEF3 (Neurofilament 3) ELISA Kit

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