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Rat GAD2 / GAD65 ELISA Kit

SKU:
RTFI00792
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
Q05683
Sensitivity:
0.094ng/ml
Range:
0.156-10ng/ml
ELISA Type:
Sandwich
Synonyms:
GAD2, GAD65, EC 4.1.1, EC 4.1.1.15, GAD65, GAD-65, GAD65MGC161605, glutamate decarboxylase 2, glutamate decarboxylase 2, pancreatic islets and brain, 65kD, glutamate decarboxylase 2, pancreatic islets and brain, 65kDa, Glutamate decarboxylase 65 kDa
Reactivity:
Rat
Research Area:
Neuroscience
€649
Frequently bought together:

Description

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Rat GAD2/GAD65 ELISA Kit

The Rat GAD2 (GAD65) ELISA Kit is a reliable and accurate tool for measuring levels of glutamate decarboxylase 2 (GAD65) in rat serum, plasma, and tissue homogenates. This kit boasts high sensitivity and specificity, ensuring precise and reproducible results for a variety of research applications.GAD65 is an important enzyme involved in the synthesis of the neurotransmitter gamma-aminobutyric acid (GABA), playing a crucial role in regulating neuronal activity in the brain.

Dysregulation of GAD65 has been implicated in conditions such as epilepsy, Parkinson's disease, and schizophrenia, making it a valuable biomarker for studying these disorders and developing potential treatments.Overall, the Rat GAD2 (GAD65) ELISA Kit provides researchers with a powerful tool for investigating the role of GAD65 in neurological disorders and advancing our understanding of brain function.

Product Name:Rat GAD2 (Glutamate Decarboxylase 2, Acid) ELISA Kit
Product Code:RTFI00792
Size:96 Assays
Target:Rat GAD2
Alias:GAD2, GAD65, EC 4.1.1, EC 4.1.1.15, GAD65, GAD-65, GAD65MGC161605, glutamate decarboxylase 2, glutamate decarboxylase 2, pancreatic islets and brain, 65kD, glutamate decarboxylase 2, pancreatic islets and brain, 65kDa, Glutamate decarboxylase 65 kDa isoform, Glutamate decarboxylase-2, pancreas, MGC161607, 65 kDa glutamic acid decarboxylase
Reactivity:Rat
Detection Method:Sandwich ELISA, Double Antibody
Sensitivity:0.094ng/ml
Range:0.156-10ng/ml
Storage:4°C for 6 months
Note:For Research Use Only
Recovery:Matrices listed below were spiked with certain level of Rat GAD2 and the recovery rates were calculated by comparing the measured value to the expected amount of Rat GAD2 in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)85-9690
EDTA plasma(n=5)91-10096
UFH plasma(n=5)93-10197
Linearity:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Rat GAD2 and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)85-104%87-103%90-104%
EDTA plasma(n=5)89-98%82-101%87-101%
UFH plasma(n=5)92-98%81-99%81-98%
Intra-Assay:CV <8%
Inter-Assay:CV <10%
Uniprot:Q05683
UniProt Protein Function:GAD2: Catalyzes the production of GABA. Homodimer. Belongs to the group II decarboxylase family.
UniProt Protein Details:

Protein type:Amino Acid Metabolism - alanine, aspartate and glutamate; Carbohydrate Metabolism - butanoate; EC 4.1.1.15; Lyase; Other Amino Acids Metabolism - beta-alanine; Other Amino Acids Metabolism - taurine and hypotaurine

Chromosomal Location of Human Ortholog: 17q12.3

Cellular Component: axon; cell junction; cytoplasm; cytoplasmic vesicle; cytosol; Golgi membrane; perinuclear region of cytoplasm; presynaptic membrane; synapse; synaptic vesicle membrane

Molecular Function:glutamate binding; glutamate decarboxylase activity; protein heterodimerization activity; pyridoxal phosphate binding

Biological Process: glutamate decarboxylation to succinate; neurotransmitter biosynthetic process; response to drug; synaptic transmission

NCBI Summary:catalyzes synthesis of gamma-Aminobutyric acid (GABA), the most widely distributed inhibitory neurotransmitter [RGD, Feb 2006]
UniProt Code:Q05683
NCBI GenInfo Identifier:6978871
NCBI Gene ID:24380
NCBI Accession:NP_036695.1
UniProt Related Accession:Q05683
Molecular Weight:65,402 Da
NCBI Full Name:glutamate decarboxylase 2
NCBI Synonym Full Names:glutamate decarboxylase 2
NCBI Official Symbol:Gad2  
NCBI Official Synonym Symbols:gad65  
NCBI Protein Information:glutamate decarboxylase 2
UniProt Protein Name:Glutamate decarboxylase 2
UniProt Synonym Protein Names:65 kDa glutamic acid decarboxylase; GAD-65; Glutamate decarboxylase 65 kDa isoform
Protein Family:Glutamate decarboxylase
UniProt Gene Name:Gad2  
StepProcedure
1. Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells!
2.Aliquot 0.1ml standard solutions into the standard wells.
3. Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well.
4. Add 0.1 ml of properly diluted sample ( Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells.
5.Seal the plate with a cover and incubate at 37°C for 90 min.
6. Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2.
7. Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall.
8.Seal the plate with a cover and incubate at 37°C for 60 min.
9. Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash.
10. Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min.
11. Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min.
12. Add 90 µL of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color.
13. Add 50 µL of Stop solution into each well and mix thoroughly. The color changes into yellow immediately.
14. Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.

Sample Type Protocol
Serum:

If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

If serum separator tubes are not being used, allow samples to clotovernight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Removeserum and assay promptly or aliquot and store the samples at-80°C. Avoid multiple freeze-thaw cycles.

Plasma: Collect plasma using EDTA or heparin as an anti-coagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.Note: Over haemolysed samples are not suitable for use with this kit.
Urine & Cerebrospinal Fluid: Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.
Cell Culture Supernatant: Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.
Cell Lysates: Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20°C.
Tissue Homogenates: The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenizein 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or-80°C.
Tissue Lysates: Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Breast Milk: Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.

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