DESCRIPTION

Synaptic vesicle membrane docking and fusion is mediated by soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNARE) complex which is located on the vesicle membrane (v-SNAREs) and the target membrane (t-SNAREs). Other than VAMP2 and STX1A, SNAP25 is another key component of SNARE core complex, and is known to be involved in regulating neurotransmitter release. Its palmitoylation domain is located in the middle of the molecule that contains four cysteine residues and mutation of the cysteines abolishes palmitoylation and its membrane binding activity. As an important presynaptic plasma membrane protein, SNAP25 has been linked to memory and learning by its effect on long term potentiation in the hippocampus, thus playing a critical role in the synaptic function of specific neuronal systems. It has been reported that SNAP25 levels were were significantly decreased in brain tissue homogenates of patients with later stages of Alzheimer's disease compared with the controls, while the levels of SNAP25 were significantly increased in cerebrospinal fluid (CSF) of the group with an Alzheimer's disease biomarker profile than in the group with a control biomarker profile.

PRINCIPLE OF THE ASSAY

A capture antibody is pre-coated onto the bottom of wells which binds to analyte of interest. A detection antibody also binds to the analyte. Horseradish peroxidase (HRP)-conjugated secondary antibody binds to the detection antibody. TMB acts as the HRP substrate and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns solution yellow. The color intensity is proportional to the quantity of bound protein which is measurable at 450 nm with the correction wavelength set at 630 nm.

PRECISION

Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested 20 times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in 24 separate assays to assess inter-assay precision.

Intra-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	20	36.9	1.1	2.9
2	20	334.6	11.2	3.3
3	20	604.7	27.9	4.6

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	24	47.1	2.9	6.0
2	24	439.0	30.3	6.9
3	24	841.1	67.0	8.0

RECOVERY

The recovery of human/mouse/rat SNAP25 spiked to three different levels throughout the range of the assay in tissue lysate.

Sample Type		Average% of Expected	Range (%)
Tissue lysate	1:250	92	81-103
	1:500	102	83-119

LINEARITY

To assess the linearity of the assay, samples were diluted with the appropriate Sample Diluent PT1-ec to produce samples with values within the dynamic range of the assay. (The samples were initially diluted 1:20)

		Tissue lysate
1:2	Average% of Expected	93
	Range (%)	85-102
1:4	Average% of Expected	111
	Range (%)	100-119
1:8	Average% of Expected	104
	Range (%)	100-112
1:16	Average% of Expected	98
	Range (%)	86-102

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human/mouse/rat SNAP25 produced at Sunlong Medical™.

Ordering & shipping

• Minimum order: 1 kit
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• Price: USD, per kit (48T / 96T options available)