Human Transcription Factor Binding To IGHM Enhancer 3 (TFE3) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to TFE3. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to TFE3. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After the TMB substrate solution is added, only those wells that contain TFE3, biotin-conjugated antibody, and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution, and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of TFE3 in the samples is then determined by comparing the O.D. of the samples to the standard curve.

For use with: Tissue homogenates and Other biological fluids.

For Research Use Only. Not for use in diagnostic procedures.

Target Information

Transcription factor that acts as a master regulator of lysosomal biogenesis and immune response (PubMed:2338243, PubMed:29146937, PubMed:30733432, PubMed:31672913). Specifically recognizes and binds E-box sequences (5'-CANNTG-3'); efficient DNA-binding requires dimerization with itself or with another MiT/TFE family member such as TFEB or MITF (By similarity). Involved in the cellular response to amino acid availability by acting downstream of MTOR: in the presence of nutrients, TFE3 phosphorylation by MTOR promotes its cytosolic retention and subsequent inactivation (PubMed:31672913). Upon starvation or lysosomal stress, inhibition of MTOR induces TFE3 dephosphorylation, resulting in nuclear localization and transcription factor activity (PubMed:31672913). Maintains the pluripotent state of embryonic stem cells by promoting the expression of genes such as ESRRB; mTOR-dependent TFE3 cytosolic retention and inactivation promotes exit from pluripotency (By similarity). Required to maintain the naive pluripotent state of hematopoietic stem cell; mTOR-dependent cytoplasmic retention of TFE3 promotes the exit of hematopoietic stem cell from pluripotency (PubMed:30733432). TFE3 activity is also involved in the inhibition of neuronal progenitor differentiation (By similarity). Acts as a positive regulator of browning of adipose tissue by promoting expression of target genes; mTOR-dependent phosphorylation promotes cytoplasmic retention of TFE3 and inhibits browning of adipose tissue (By similarity). In association with TFEB, activates the expression of CD40L in T-cells, thereby playing a role in T-cell-dependent antibody responses in activated CD4(+) T-cells and thymus-dependent humoral immunity (By similarity). Specifically recognizes the MUE3 box, a subset of E-boxes, present in the immunoglobulin enhancer (PubMed:2338243). It also binds very well to a USF/MLTF site (PubMed:2338243). May regulate lysosomal positioning in response to nutrient deprivation by promoting the expression of PIP4P1 (PubMed:29146937).

GENE ID	7030
SWISS PROT	P19532
SYNONYMS	RCCP2; TFEA; BHLHE33; Transcription Tactor E Family Member A; Class E basic helix-loop-helix protein 33; Transcription factor E3

Materials Supplied

Kit Components	96 Wells Quantity/Size
Pre-coated, ready-to-use 96-well strip plate	1 plate
Plate sealer for 96 wells	2
Standard	2 tubes
Diluent buffer	1 bottle
Detection Reagent A	1 bottle
Detection Reagent B	1 bottle
TMB Substrate	1 tube
Stop Solution	1 tube
Wash Buffer (30X concentrate)	1 tube
Product data sheet	1 copy

Storage

Storage

The TMB Substrate, Wash Buffer (30X concentrate), and the Stop Solution should be stored at 4°C upon receipt, while the other items should be stored at -20°C.

Performance Characteristics

REPEATABILITY	Intra-assay Precision (Precision within an assay): 3 samples with low, middle, and high-level TFE3 were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level TFE3 were tested on 3 different plates, with 8 replicates in each plate. CV(%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%
SENSITIVITY	The minimum detectable dose was 0.118ng/mL.
ASSAY RANGE	0.312–20ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of TFE3. No significant cross-reactivity or interference between TFE3 and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between TFE3 and all analogs, therefore, cross reactivity may still exist.