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Human GABA / Gamma-aminobutyric acid ELISA Kit

SKU:
HUFI02481
Product Type:
ELISA Kit
Size:
96 Assays
Sensitivity:
18.75pg/ml
Range:
31.25-2000pg/ml
ELISA Type:
Competitive
Synonyms:
GABA
Reactivity:
Human
€599
Frequently bought together:

Description

Human GABA / Gamma-aminobutyric acid ELISA Kit

GABA, a vital neurotransmitter in the central nervous system, plays a crucial role in regulating brain activity and promoting a balanced mental state. By binding to specific receptors in the brain, GABA helps to balance and control the excitability of nerve cells, promoting a state of calm and relaxation. The Assay Genie Human GABA / Gamma-aminobutyric acid ELISA Kit is a highly sensitive assay for the quantitative measurement of FSH in serum, plasma, cell and tissue lysates.

system_update_alt Datasheet system_update_alt MSDS

Key Features

Save Time Pre-coated 96 well plate
Quick Start Kit includes all necessary reagents
Publication Ready Reproducible and reliable results

Overview

Product Name:

Human GABA/ Gamma-aminobutyric acid ELISA Kit

Product Code:

HUFI02481

Size:

96 Assays

Alias:

GABA

Detection Method:

Competitive ELISA, Coated with Antibody

Application:

This immunoassay kit allows for the in vitro quantitative determination of Human GABA concentrations in serum plasma and other biological fluids.

Sensitivity:

18.75pg/ml

Range:

31.25-2000pg/ml

Storage:

4°C for 6 months

Note:

For Research Use Only

Additional Information

Recovery

Matrices listed below were spiked with certain level of Human GABA and the recovery rates were calculated by comparing the measured value to the expected amount of Human GABA in samples.

Matrix

Recovery Range (%)

Average (%)

serum(n=5)

98-101

99

EDTA plasma(n=5)

85-99

93

UFH plasma(n=5)

92-96

94

Linearity:

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human GABA and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Sample

1:2

1:4

1:8

Text

86-102%

86-97%

85-105%

Text

86-97%

82-99%

82-98%

Text

83-100%

86-100%

80-99%

Kit Components

Component Quantity Storage

ELISA Microplate (Dismountable)

8x12 strips

4°C for 6 months

Lyophilized Standard

2

4°C/ -20°C

Sample/Standard Dlution Buffer

20ml

4°C

Biotin-labeled Antibody (Concentrated)

120ul

4°C (Protection from light)

Antibody Dilution Buffer

10ml

4°C

HRP-Streptavidin Conjugate (SABC)

120ul

4°C (Protect from light)

SABC Dilution Buffer

10ml

4°C

TMB Substrate

10ml

4°C (Protection from light)

Stop Solution

10ml

4°C

Wash Buffer (25X)

30ml

4°C

Plate Sealer

5

-

Other materials required:

  • Microplate reader with 450 nm wavelength filter
  • Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
  • Incubator
  • Deionized or distilled water
  • Absorbent paper
  • Buffer resevoir

Protocol

*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.

Equilibrate the TMB substrate for at least 30 min at 37°C beforeuse. When diluting samples and reagents, they must be mixed completely andevenly. It is recommended to plot a standard curve for each test.

Step Procedure

1.

Set standard, test sample and control (zero) wells on the pre-coatedplate respectively, and then, record their positions. It isrecommended to measure each standard and sample in duplicate. Washplate 2 times before adding standard, sample and control (zero) wells!

2.

Add Sample and Biotin-detection antibody: Add 50µL of Standard, Blank or Sample per well. The blankwell is added with Sample Dilution Buffer. Immediately add 50 µL of biotin-labelled antibody workingsolution to each well. Cover with the plate sealer provided. Gently tap the plate to ensure thoroughmixing. Incubate for 45 minutes at 37°C. (Solutions are added to the bottom of micro-ELISA platewell, avoid touching plate walls and foaming).

3.

Wash: Aspirate each well and wash, repeating the process three timesWash by filling each well with Wash Buffer (approximately 350µL)using a squirt bottle, multi-channel pipette, manifold dispenser orautomated washer. Complete removal of liquid at each step is essentialto good performance. After the last wash, remove any remaining WashBuffer by aspirating or decanting. Invert the plate and pat it againstthick clean absorbent paper.

4.

HRP-Streptavidin Conjugate(SABC): Add 100µL of SABC workingsolution to each well. Cover with a new Plate sealer. Incubate for30minutes at 37°C.

5.

Wash: Repeat the aspiration/wash process for five times.

6.

TMB Substrate: Add 90µL of TMB Substrate to each well. Coverwith a new Plate sealer. Incubate for about 10-20 minutes at 37°C.Protect from light. The reaction time can be shortened or extendedaccording to the actual color change, but not more than 30minutes.When apparent gradient appeared in standard wells, you can terminatethe reaction.

7.

Stop: Add 50µL of Stop Solution to each well. Color turn toyellow immediately. The adding order of stop solution should be as thesame as the substrate solution.

8.

OD Measurement: Determine the optical density (OD Value) of each wellat once, using a microplate reader set to 450 nm. You should open themicroplate reader ahead, preheat the instrument, and set the testing parameters.

Sample Type

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 clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum 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 anticoagulant. 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 & homogenize in 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.

GABA Background

GABA Background

Gamma-aminobutyric acid (GABA) is a necessary amino acid that serves as the brain's main inhibitory neurotransmitter and has a sizable inhibitory effect on the spinal cord. It attaches to post-synaptic GABA receptors in the synapses between neurons, where it performs its main action. This binding mechanism alters ion channels, causing the cell to become hyperpolarized and thereby blocking the transmission of an action potential. It is impossible to overestimate GABA's clinical importance given that several neurologic and psychiatric illnesses have been linked to abnormalities in GABA signaling. Numerous pharmaceutical treatments for neurological, psychiatric, and anaesthetic conditions are predicated on modulating GABA signaling. 

Role of GABA

Its primary function is to regulate neuronal activity and maintain a delicate balance between excitation and inhibition. By binding to GABA receptors on post-synaptic neurons, GABA hyperpolarizes the cell membrane, reducing the likelihood of an action potential being triggered. This inhibitory effect helps to modulate and dampen excessive neuronal firing, promoting a state of calm and relaxation. GABA's role in regulating the overall excitability of the brain is vital for maintaining cognitive functions, emotional stability, and motor control. Imbalances in GABA levels or dysfunction in GABAergic signaling have been associated with a range of neurological conditions, such as anxiety disorders, epilepsy, insomnia, and mood disorders. Understanding the role of GABA is essential for unravelling the intricate mechanisms underlying brain function and developing targeted therapies for various neurological disorders.

GABA Receptors

These receptors are protein structures located on the surface of neurons and are divided into two main types: GABAA and GABAB receptors. GABAA receptors are ion channels that, when activated by GABA binding, allow chloride ions to enter the cell, leading to hyperpolarization and inhibition of neuronal activity. They are responsible for fast inhibitory neurotransmission in the brain. GABAB receptors, on the other hand, are G-protein coupled receptors that exert a slower inhibitory effect by activating intracellular signaling pathways. The significance of GABA receptors cannot be overstated, as they are involved in modulating the overall excitability of the brain and spinal cord.

GABA Cycle

GABA is derived from glutamate, which itself originates from alpha-ketoglutarate. The Glutamate-GABA-glutamine cycle is a complex multicellular process responsible for the synthesis and processing of glutamate, glutamine, and GABA. Within the presynaptic neuron, the enzyme glutamate decarboxylase (GAD) utilizes pyridoxal phosphate (vitamin B6) as a co-factor to convert glutamate into GABA. Once released into the synaptic cleft, GABA can bind to GABA receptors, be recycled within the presynaptic neuron, or be taken up by astrocytes for degradation. Inside astrocytes, GABA is broken down by GABA-transaminase (GABA-T) into succinic semialdehyde (SSA). Subsequently, succinic semialdehyde is transformed into succinate-by-succinate semialdehyde dehydrogenase (SSADH). Succinate then enters the tricyclic acid (TCA) cycle and converts back into alpha-ketoglutarate, which ultimately becomes glutamate. The enzyme glutamine synthase converts glutamate into glutamine, facilitating its transportation from astrocytes to presynaptic neurons. Finally, within the presynaptic neuron, phosphate-activated glutaminase converts glutamine back into glutamate, thereby restarting the Glutamate-GABA-glutamine cycle.

GABA FAQs

What is the GABA ELISA kit used for?

The Human GABA ELISA Kit is specifically designed for the quantitative detection of Gamma-Aminobutyric Acid (GABA) in human samples. GABA is an important neurotransmitter that plays a crucial role in regulating neuronal excitability and maintaining the balance between excitation and inhibition in the central nervous system. This kit provides researchers and clinicians with a reliable and efficient tool to measure GABA levels, facilitating studies related to neurobiology, neurological disorders, and psychiatric conditions.

What are the advantages of using the GABA ELISA Kit?

The Human GABA ELISA Kit offers several advantages. Firstly, it provides high sensitivity and specificity, ensuring accurate and precise quantification of GABA levels in human samples. The kit also offers a wide dynamic range, allowing for the detection of both low and high concentrations of GABA. Additionally, it is user-friendly, with a straightforward protocol and short assay time. The kit provides reliable and reproducible results, enabling researchers to obtain valuable insights into GABA-related research areas.

What sample types are compatible with GABA ELISA kit?

The Human GABA ELISA Kit is compatible with various sample types, including human plasma, serum, cerebrospinal fluid (CSF), and tissue homogenates. The kit provides flexibility in sample selection, allowing researchers to analyze GABA levels in different biological matrices relevant to their specific research interests.

What are the storage requirements for GABA ELISA kit?

The GABA ELISA Kit components should be stored according to the instructions provided in the kit manual. Generally, it is recommended to store the kit components at the recommended temperature to ensure their stability and optimal performance.

What should I do if my assay results are not optimal?

If you encounter any issues or have suboptimal assay results, we recommend contacting our dedicated support team for assistance. They will be available to provide troubleshooting guidance, answer your questions, and ensure you achieve the best possible results with the GABA ELISA Kit.