Description
Human GLUT4 (Glucose Transporter 4) ELISA Kit
The Human GLUT4 (Glucose Transporter 4) ELISA Kit is specifically designed for the precise measurement of GLUT4 levels in human serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit delivers accurate and consistent results, making it a valuable tool for various research endeavors.GLUT4 is a key protein involved in glucose transport, playing a vital role in regulating blood sugar levels and insulin sensitivity. Dysregulation of GLUT4 function is implicated in conditions like diabetes and obesity, underscoring its importance as a biomarker for studying metabolic diseases and developing targeted treatments.
Whether investigating metabolic disorders or exploring the intricacies of glucose metabolism, the Human GLUT4 ELISA Kit offers researchers a reliable and efficient way to quantify GLUT4 levels and further our understanding of these critical physiological processes.
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Human |
| Detection Method: | Colormetric |
| Detection Range: | 0.31-20 ng/mL |
| Sensitivity: | 0.19 ng/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Human GLUT4 in samples. No significant cross-reactivity or interference between Human GLUT4 and analogues was observed. |
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human GLUT4. Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human GLUT4 and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human GLUT4, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human GLUT4. The concentration of Human GLUT4 in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | GLUT4: an integral membrane facilitative glucose transporter. One of 13 members of the human equilibrative glucose transport protein family. Its surface expression is regulated by insulin. Has two internalization sequences, a dileucine repeat present in the C-terminus and a FxxY motif in the amino-terminal end. Associates with an intracellular tubulo-vesicular compartment under low plasma conditions. The binding of insulin to the insulin receptor (a typosine kinase) leads to the rapid translocation of GLUT4 to the cell surface, increasing cellular glucose transport activity. Its translocation to the plasma membrane is also stimulated by exercise, but independent from insulin signaling. This insulin-independent pathway may be regulated by AMPK. |
| UniProt Protein Details: | Protein type:Transporter, SLC family; Transporter; Membrane protein, multi-pass; Membrane protein, integral Chromosomal Location of Human Ortholog: 17p13 Cellular Component: multivesicular body; trans-Golgi network transport vesicle; membrane; clathrin-coated vesicle; integral to plasma membrane; perinuclear region of cytoplasm; T-tubule; plasma membrane; endomembrane system; coated pit; vesicle membrane; cytosol; lipid raft; external side of plasma membrane Molecular Function:D-glucose transmembrane transporter activity; protein binding; glucose transmembrane transporter activity Biological Process: amylopectin biosynthetic process; response to ethanol; cellular response to insulin stimulus; glucose import; hexose transport; carbohydrate metabolic process; brown fat cell differentiation; pathogenesis; glucose transport; glucose homeostasis; transmembrane transport Disease: Diabetes Mellitus, Noninsulin-dependent |
| NCBI Summary: | This gene is a member of the solute carrier family 2 (facilitated glucose transporter) family and encodes a protein that functions as an insulin-regulated facilitative glucose transporter. In the absence of insulin, this integral membrane protein is sequestered within the cells of muscle and adipose tissue. Within minutes of insulin stimulation, the protein moves to the cell surface and begins to transport glucose across the cell membrane. Mutations in this gene have been associated with noninsulin-dependent diabetes mellitus (NIDDM). [provided by RefSeq, Jul 2008] |
| UniProt Code: | P14672 |
| NCBI GenInfo Identifier: | 121761 |
| NCBI Gene ID: | 6517 |
| NCBI Accession: | P14672. 1 |
| UniProt Secondary Accession: | P14672,Q05BQ3, Q14CX2, |
| UniProt Related Accession: | P14672 |
| Molecular Weight: | 509 |
| NCBI Full Name: | Solute carrier family 2, facilitated glucose transporter member 4 |
| NCBI Synonym Full Names: | solute carrier family 2 (facilitated glucose transporter), member 4 |
| NCBI Official Symbol: | SLC2A4 |
| NCBI Official Synonym Symbols: | GLUT4 |
| NCBI Protein Information: | solute carrier family 2, facilitated glucose transporter member 4; GLUT-4; insulin-responsive glucose transporter type 4; glucose transporter type 4, insulin-responsive |
| UniProt Protein Name: | Solute carrier family 2, facilitated glucose transporter member 4 |
| UniProt Synonym Protein Names: | Glucose transporter type 4, insulin-responsive; GLUT-4 |
| UniProt Gene Name: | SLC2A4 |
| UniProt Entry Name: | GTR4_HUMAN |
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
| Concentration (ng/mL) | O.D | Average | Corrected |
| 20 | 2.385 2.429 | 2.407 | 2.348 |
| 10 | 1.598 1.624 | 1.611 | 1.552 |
| 5 | 0.983 0.967 | 0.975 | 0.916 |
| 2.5 | 0.522 0.53 | 0.526 | 0.467 |
| 1.25 | 0.262 0.246 | 0.254 | 0.195 |
| 0.63 | 0.176 0.154 | 0.165 | 0.106 |
| 0.31 | 0.111 0.113 | 0.112 | 0.053 |
| 0 | 0.05 0.068 | 0.059 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human GLUT4 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human GLUT4 were tested on 3 different plates, 20 replicates in each plate.
| Intra-assay Precision | Inter-assay Precision | |||||
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (ng/mL) | 1.07 | 3.17 | 8.74 | 1.13 | 3.12 | 9.09 |
| Standard deviation | 0.05 | 0.17 | 0.48 | 0.06 | 0.14 | 0.42 |
| C V (%) | 4.67 | 5.36 | 5.49 | 5.31 | 4.49 | 4.62 |
Recovery
The recovery of Human GLUT4 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
| Sample Type | Range (%) | Average Recovery (%) |
| Serum (n=5) | 94-110 | 101 |
| EDTA plasma (n=5) | 87-101 | 94 |
| Cell culture media (n=5) | 94-109 | 101 |
Linearity
Samples were spiked with high concentrations of Human GLUT4 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
| Serum (n=5) | EDTA plasma (n=5) | Cell culture media (n=5) | ||
| 1:2 | Range (%) | 96-112 | 99-112 | 88-101 |
| Average (%) | 103 | 106 | 94 | |
| 1:4 | Range (%) | 86-102 | 82-96 | 81-92 |
| Average (%) | 93 | 89 | 87 | |
| 1:8 | Range (%) | 87-100 | 82-95 | 86-98 |
| Average (%) | 94 | 87 | 92 | |
| 1:16 | Range (%) | 92-104 | 81-93 | 81-96 |
| Average (%) | 98 | 86 | 88 |
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
| Item | Specifications | Storage |
| Micro ELISA Plate(Dismountable) | 8 wells ×12 strips | -20°C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100×) | 1 vial, 120 µL | |
| Concentrated HRP Conjugate (100×) | 1 vial, 120 µL | -20°C(shading light), 6 months |
| Reference Standard & Sample Diluent | 1 vial, 20 mL | 4°C, 6 months |
| Biotinylated Detection Ab Diluent | 1 vial, 14 mL | |
| HRP Conjugate Diluent | 1 vial, 14 mL | |
| Concentrated Wash Buffer (25×) | 1 vial, 30 mL | |
| Substrate Reagent | 1 vial, 10 mL | 4°C(shading light) |
| Stop Solution | 1 vial, 10 mL | 4°C |
| Plate Sealer | 5 pieces | |
| Product Description | 1 copy | |
| Certificate of Analysis | 1 copy |
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37°C.
- Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
