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Human ATP5G3 (ATP synthase lipid binding protein, mitochondrial) ELISA Kit (HUFI07679)

SKU:
HUFI07679
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P48201
Sensitivity:
9.375pg/ml
Range:
15.625-1000pg/ml
ELISA Type:
Sandwich
Synonyms:
ATP5G3
Reactivity:
Human
€599
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Description

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Human ATP5G3 (ATP synthase lipid binding protein, mitochondrial) ELISA Kit (HUFI07679)

The Human ATP5G3 (ATP Synthase Lipid Binding Protein) Mitochondrial ELISA Kit is specifically designed for the accurate detection of ATP5G3 levels in human serum, plasma, and cell culture supernatants. This advanced kit offers high sensitivity and specificity, ensuring precise and reliable results for a variety of research applications.ATP5G3 is a key protein involved in mitochondrial ATP synthesis, playing a vital role in cellular energy production. Dysregulation of ATP5G3 has been implicated in various diseases, including metabolic disorders and neurodegenerative conditions.

Therefore, this ELISA kit serves as a valuable tool for studying ATP5G3 levels in different biological samples and exploring its potential implications in disease development and progression.Overall, the Human ATP5G3 Mitochondrial ELISA Kit from AssayGenie provides researchers with a powerful means to investigate the role of ATP5G3 in cellular metabolism and disease pathology, paving the way for potential therapeutic interventions.

Product Name:Human ATP5G3 (ATP synthase lipid binding protein, mitochondrial) ELISA Kit
Product Code:HUFI07679
Size:96 Assays
Alias:ATP5G3 ELISA Kit
Detection method:Sandwich ELISA, Double Antibody
Application:This immunoassay kit allows for the in vitro quantitative determination of Human ATP5G3 (ATP synthase lipid binding protein, mitochondrial) concentrations in serum plasma and other biological fluids.
Sensitivity:< 9.375pg/ml
Range:15.625-1000pg/ml
Storage:4°C for 6 months
Note:For Research Use Only
Recovery:Matrices listed below were spiked with certain level of Human ATP5G3 (ATP synthase lipid binding protein, mitochondrial) and the recovery rates were calculated by comparing the measured value to the expected amount of Human ATP5G3 (ATP synthase lipid binding protein, mitochondrial) in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)88-10296
EDTA plasma(n=5)89-10496
heparin plasma(n=5)85-10396
Linearity:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human ATP5G3 (ATP synthase lipid binding protein, mitochondrial) and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)86-103%91-103%90-103%
EDTA plasma(n=5)83-101%83-101%84-98%
UFH plasma(n=5)80-97%82-99%80-98%
CV(%):Intra-Assay: CV<8%
Inter-Assay: CV<10%
ComponentQuantityStorage
ELISA Microplate (Dismountable)8×12 strips4°C for 6 months
Lyophilized Standard24°C/-20°C
Sample/Standard Dilution Buffer20ml4°C
Biotin-labeled Antibody(Concentrated)120ul4°C (Protect from light)
Antibody Dilution Buffer10ml4°C
HRP-Streptavidin Conjugate(SABC)120ul4°C (Protect from light)
SABC Dilution Buffer10ml4°C
TMB Substrate10ml4°C (Protect from light)
Stop Solution10ml4°C
Wash Buffer(25X)30ml4°C
Plate Sealer5-

Other materials and equipment 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
UniProt Protein Function:ATP5G3: Mitochondrial membrane ATP synthase (F(1)F(0) ATP synthase or Complex V) produces ATP from ADP in the presence of a proton gradient across the membrane which is generated by electron transport complexes of the respiratory chain. F-type ATPases consist of two structural domains, F(1) - containing the extramembraneous catalytic core and F(0) - containing the membrane proton channel, linked together by a central stalk and a peripheral stalk. During catalysis, ATP synthesis in the catalytic domain of F(1) is coupled via a rotary mechanism of the central stalk subunits to proton translocation. Part of the complex F(0) domain. A homomeric c-ring of probably 10 subunits is part of the complex rotary element. Belongs to the ATPase C chain family.
UniProt Protein Details:Protein type:Energy Metabolism - oxidative phosphorylation; Membrane protein, integral; Membrane protein, multi-pass

Chromosomal Location of Human Ortholog: 2q31.1

Cellular Component: mitochondrial outer membrane; mitochondrial proton-transporting ATP synthase complex

Biological Process: ATP biosynthetic process; mitochondrial ATP synthesis coupled proton transport

NCBI Summary:This gene encodes a subunit of mitochondrial ATP synthase. Mitochondrial ATP synthase catalyzes ATP synthesis, utilizing an electrochemical gradient of protons across the inner membrane during oxidative phosphorylation. ATP synthase is composed of two linked multi-subunit complexes: the soluble catalytic core, F1, and the membrane-spanning component, Fo, comprising the proton channel. The catalytic portion of mitochondrial ATP synthase consists of 5 different subunits (alpha, beta, gamma, delta, and epsilon) assembled with a stoichiometry of 3 alpha, 3 beta, and a single representative of the other 3. The proton channel seems to have nine subunits (a, b, c, d, e, f, g, F6 and 8). This gene is one of three genes that encode subunit c of the proton channel. Each of the three genes have distinct mitochondrial import sequences but encode the identical mature protein. Alternatively spliced transcript variants encoding different proteins have been identified. [provided by RefSeq, Jun 2010]
UniProt Code:P48201
NCBI GenInfo Identifier:1352048
NCBI Gene ID:518
NCBI Accession:P48201.1
UniProt Secondary Accession:P48201,Q4ZFX7, B2R4Z0, D3DPF0,
UniProt Related Accession:P48201
Molecular Weight:14,693 Da
NCBI Full Name:ATP synthase F(0) complex subunit C3, mitochondrial
NCBI Synonym Full Names:ATP synthase, H+ transporting, mitochondrial Fo complex subunit C3 (subunit 9)
NCBI Official Symbol:ATP5G3
NCBI Official Synonym Symbols:P3
NCBI Protein Information:ATP synthase F(0) complex subunit C3, mitochondrial
UniProt Protein Name:ATP synthase F(0) complex subunit C3, mitochondrial
UniProt Synonym Protein Names:ATP synthase lipid-binding protein; ATP synthase proteolipid P3; ATP synthase proton-transporting mitochondrial F(0) complex subunit C3; ATPase protein 9; ATPase subunit c
UniProt Gene Name:ATP5G3
UniProt Entry Name:AT5G3_HUMAN

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

Before adding to wells, equilibrate the SABC working solution and TMB substrate for at least 30 min at 37 °C. When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.

StepProtocol
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 TypeProtocol
SerumIf 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.

PlasmaCollect 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 FluidCollect 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 supernatantCollect 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 lysatesSolubilize 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 homogenatesThe 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 lysatesRinse 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 MilkCollect 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.
Antibodies
Anti-ATP5G3 Antibody (CAB18278)
ATP5G3 Antibody (PACO04574)
ATP5G3 Antibody (PACO23389)

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