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Human Hexosaminidase B / HEXB ELISA Kit

SKU:
HUFI02055
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P07686
Sensitivity:
0.938IU/ml
Range:
1.563-100IU/ml
ELISA Type:
Sandwich
Synonyms:
HEXB, ENC-1AS, N-acetyl-beta-glucosaminidase subunit beta, Beta-N-acetylhexosaminidase subunit beta, Hexosaminidase subunit B, Cervical cancer proto-oncogene 7 protein, HCC-7, ENC-1AS, HCC-7, HEL-248, beta-hexosaminidase subunit beta, Beta-N-acetylhe
Reactivity:
Human
Research Area:
Signal Transduction
€599
Frequently bought together:

Description

Human Hexosaminidase B/HEXB ELISA Kit

The Human HEXB (Hexosaminidase B) ELISA Kit is a powerful tool for the accurate measurement of HEXB levels in human samples such as serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit provides reliable and reproducible results, making it ideal for a variety of research purposes.HEXB is an important enzyme that plays a key role in the breakdown of glycolipids and glycoproteins in lysosomes.

Mutations in the HEXB gene can lead to Tay-Sachs disease, a rare and devastating genetic disorder. Therefore, measuring HEXB levels can be valuable in understanding lysosomal storage disorders and developing potential treatments.Get your Human HEXB ELISA Kit today and enhance your research on lysosomal storage disorders and related conditions.

Product Name:Human Hexosaminidase B / HEXB ELISA Kit
Product Code:HUFI02055
Size:96 Assays
Alias:HEXB, ENC-1AS, N-acetyl-beta-glucosaminidase subunit beta, Beta-N-acetylhexosaminidase subunit beta, Hexosaminidase subunit B, Cervical cancer proto-oncogene 7 protein, HCC-7, ENC-1AS, HCC-7, HEL-248, beta-hexosaminidase subunit beta, Beta-N-acetylhexosaminidase subunit beta, Cervical cancer proto-oncogene 7 protein, hexosaminidase B, beta polypeptide, Hexosaminidase subunit B, N-acetyl-beta-glucosaminidase subunit beta
Detection method:Sandwich ELISA, Double Antibody
Application:This immunoassay kit allows for the in vitro quantitative determination of Human HEXB concentrations in serum plasma and other biological fluids.
Sensitivity:0.938IU/ml
Range:1.56-100IU/ml
Storage:4°C for 6 months
Note:For Research Use Only
Recovery:Matrices listed below were spiked with certain level of Human HEXB and the recovery rates were calculated by comparing the measured value to the expected amount of Human HEXB in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)85-10596
EDTA plasma(n=5)85-10197
UFH plasma(n=5)85-9790
Linearity:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human HEXB and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)88-103%87-100%89-103%
EDTA plasma(n=5)89-101%85-98%86-97%
UFH plasma(n=5)88-100%81-96%83-93%
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
UniprotP07686
UniProt Protein Function:HEXB: Responsible for the degradation of GM2 gangliosides, and a variety of other molecules containing terminal N-acetyl hexosamines, in the brain and other tissues. Defects in HEXB are the cause of GM2-gangliosidosis type 2 (GM2G2); also known as Sandhoff disease. GM2- gangliosidosis is an autosomal recessive lysosomal storage disease marked by the accumulation of GM2 gangliosides in the neuronal cells. GM2G2 is clinically indistinguishable from GM2- gangliosidosis type 1, presenting startle reactions, early blindness, progressive motor and mental deterioration, macrocephaly and cherry-red spots on the macula. Belongs to the glycosyl hydrolase 20 family.
UniProt Protein Details:Protein type:Carbohydrate Metabolism - amino sugar and nucleotide sugar; EC 3.2.1.52; Glycan Metabolism - glycosaminoglycan degradation; Glycan Metabolism - glycosphingolipid biosynthesis - ganglio series; Glycan Metabolism - glycosphingolipid biosynthesis - globo series; Glycan Metabolism - other glycan degradation; Hydrolase

Chromosomal Location of Human Ortholog: 5q13.3

Cellular Component: acrosomal vesicle; azurophil granule; extracellular region; lysosomal lumen; membrane

Molecular Function:acetylglucosaminyltransferase activity; beta-N-acetylhexosaminidase activity; protein heterodimerization activity; protein homodimerization activity

Biological Process: astrocyte cell migration; cellular calcium ion homeostasis; cellular protein metabolic process; chondroitin sulfate catabolic process; ganglioside catabolic process; glycosphingolipid metabolic process; hyaluronan catabolic process; keratan sulfate catabolic process; lipid storage; locomotory behavior; lysosome organization and biogenesis; male courtship behavior; myelination; neuromuscular process controlling balance; neutrophil degranulation; oligosaccharide catabolic process; oogenesis; penetration of zona pellucida; phospholipid biosynthetic process; positive regulation of transcription from RNA polymerase II promoter; regulation of cell shape; sensory perception of sound; skeletal system development

Disease: Sandhoff Disease

NCBI Summary:Hexosaminidase B is the beta subunit of the lysosomal enzyme beta-hexosaminidase that, together with the cofactor GM2 activator protein, catalyzes the degradation of the ganglioside GM2, and other molecules containing terminal N-acetyl hexosamines. Beta-hexosaminidase is composed of two subunits, alpha and beta, which are encoded by separate genes. Both beta-hexosaminidase alpha and beta subunits are members of family 20 of glycosyl hydrolases. Mutations in the alpha or beta subunit genes lead to an accumulation of GM2 ganglioside in neurons and neurodegenerative disorders termed the GM2 gangliosidoses. Beta subunit gene mutations lead to Sandhoff disease (GM2-gangliosidosis type II). Alternatively spliced transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, May 2014]
UniProt Code:P07686
NCBI GenInfo Identifier:4504373
NCBI Gene ID:3074
NCBI Accession:NP_000512.1
UniProt Related Accession:P07686
Molecular Weight:
NCBI Full Name:beta-hexosaminidase subunit beta isoform 1 preproprotein
NCBI Synonym Full Names:hexosaminidase subunit beta
NCBI Official Symbol:HEXB  
NCBI Official Synonym Symbols:ENC-1AS; HEL-248; HEL-S-111  
NCBI Protein Information:beta-hexosaminidase subunit beta
UniProt Protein Name:Beta-hexosaminidase subunit beta
UniProt Synonym Protein Names:Beta-N-acetylhexosaminidase subunit beta; Hexosaminidase subunit B
Protein Family:Beta-hexosaminidase
UniProt Gene Name:HEXB  

*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.

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