null

Human Carbonic anhydrase 4 / CA4 ELISA Kit

SKU:
HUFI00994
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P22748
Sensitivity:
0.469ng/ml
Range:
0.781-50ng/ml
ELISA Type:
Sandwich
Synonyms:
CA4, Carbonic Anhydrase IV, CAIV, CA-IV, Car4, RP17, Carbonate dehydRatase IV
Reactivity:
Human
Research Area:
Cell Biology
€499
Frequently bought together:

Description

system_update_altDatasheetsystem_update_altMSDS

Human Carbonic anhydrase 4/CA4 ELISA Kit

The Human Carbonic Anhydrase 4 (CA4) ELISA Kit is a highly sensitive and specific assay designed for the accurate detection of CA4 levels in human serum, plasma, and cell culture supernatants. This kit provides reliable and reproducible results, making it ideal for a wide range of research applications.Carbonic anhydrase 4 is an important enzyme that plays a key role in maintaining acid-base balance in the body. It is involved in processes such as acidification of the urine and secretion of stomach acid.

Abnormal levels of CA4 have been associated with various diseases including renal tubular acidosis and osteopetrosis, making it a valuable biomarker for studying these conditions and developing potential therapies.Overall, the Human Carbonic Anhydrase 4 ELISA Kit is a powerful tool for researchers looking to investigate the role of CA4 in health and disease, with its high sensitivity and specificity ensuring accurate and reliable results.

Product Name:Human Carbonic anhydrase 4 / CA4 ELISA Kit
Product Code:HUFI00994
Size:96 Assays
Alias:CA4, Carbonic Anhydrase IV, CAIV, CA-IV, Car4, RP17, Carbonate dehydRatase IV
Detection method:Sandwich ELISA, Double Antibody
Application:This immunoassay kit allows for the in vitro quantitative determination of Human CA4 concentrations in serum plasma and other biological fluids.
Sensitivity:0.469ng/ml
Range:0.781-50ng/ml
Storage:4°C for 6 months
Note:For Research Use Only
Recovery:Matrices listed below were spiked with certain level of Human CA4 and the recovery rates were calculated by comparing the measured value to the expected amount of Human CA4 in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)85-10192
EDTA plasma(n=5)85-10096
UFH plasma(n=5)93-10498
Linearity:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human CA4 and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)86-104%86-99%88-99%
EDTA plasma(n=5)82-101%87-99%83-99%
UFH plasma(n=5)86-93%82-98%81-93%
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
UniprotP22748
UniProt Protein Function:CA4: Reversible hydration of carbon dioxide. May stimulate the sodium/bicarbonate transporter activity of SLC4A4 that acts in pH homeostasis. It is essential for acid overload removal from the retina and retina epithelium, and acid release in the choriocapillaris in the choroid. Defects in CA4 are the cause of retinitis pigmentosa type 17 (RP17). RP leads to degeneration of retinal photoreceptor cells. Patients typically have night vision blindness and loss of midperipheral visual field. As their condition progresses, they lose their far peripheral visual field and eventually central vision as well. RP17 inheritance is autosomal dominant. Defective acid overload removal from retina and retinal epithelium, due to mutant CA4, is responsible for photoreceptor degeneration, indicating that impaired pH homeostasis is the most likely cause underlying the RP17 phenotype. Belongs to the alpha-carbonic anhydrase family.
UniProt Protein Details:Protein type:Lyase; Energy Metabolism - nitrogen; Membrane protein, GPI anchor; EC 4.2.1.1

Chromosomal Location of Human Ortholog: 17q23

Cellular Component: Golgi apparatus; cell surface; sarcoplasmic reticulum; rough endoplasmic reticulum; basolateral plasma membrane; brush border membrane; secretory granule membrane; ER-Golgi intermediate compartment; trans-Golgi network; anchored to external side of plasma membrane; membrane; perinuclear region of cytoplasm; apical plasma membrane; plasma membrane; sarcolemma

Molecular Function:carbonate dehydratase activity; protein binding; zinc ion binding

Biological Process: response to drug; response to steroid hormone stimulus; bicarbonate transport; organ development; one-carbon compound metabolic process

Disease: Retinitis Pigmentosa 17

NCBI Summary:Carbonic anhydrases (CAs) are a large family of zinc metalloenzymes that catalyze the reversible hydration of carbon dioxide. They participate in a variety of biological processes, including respiration, calcification, acid-base balance, bone resorption, and the formation of aqueous humor, cerebrospinal fluid, saliva, and gastric acid. They show extensive diversity in tissue distribution and in their subcellular localization. This gene encodes a glycosylphosphatidyl-inositol-anchored membrane isozyme expressed on the luminal surfaces of pulmonary (and certain other) capillaries and proximal renal tubules. Its exact function is not known; however, it may have a role in inherited renal abnormalities of bicarbonate transport. [provided by RefSeq, Jul 2008]
UniProt Code:P22748
NCBI GenInfo Identifier:115465
NCBI Gene ID:762
NCBI Accession:P22748.2
UniProt Secondary Accession:P22748,Q64444, P48284,
UniProt Related Accession:P22748
Molecular Weight:312
NCBI Full Name:Carbonic anhydrase 4
NCBI Synonym Full Names:carbonic anhydrase IV
NCBI Official Symbol:CA4
NCBI Official Synonym Symbols:CAIV; Car4; RP17
NCBI Protein Information:carbonic anhydrase 4; CA-IV; carbonic dehydratase IV; carbonate dehydratase IV
UniProt Protein Name:Carbonic anhydrase 4
UniProt Synonym Protein Names:Carbonate dehydratase IV; Carbonic anhydrase IV; CA-IV
Protein Family:Carbonic anhydrase
UniProt Gene Name:CA4
UniProt Entry Name:CAH4_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.

Related Products