The Mouse Nuclear Receptor ROR Gamma (RORC) ELISA Kit is specifically designed for the accurate detection of RORC levels in mouse serum, plasma, and cell culture supernatants. This kit offers high sensitivity and specificity, ensuring reliable and reproducible results for a variety of research applications.RORC is a nuclear receptor that plays a crucial role in the regulation of immune responses and the development of T cells. Dysregulation of RORC has been implicated in various autoimmune diseases, making it an important biomarker for studying immune system function and developing targeted therapies.
With its advanced features and high-quality components, the Mouse Nuclear Receptor ROR Gamma (RORC) ELISA Kit is an essential tool for researchers seeking to understand the role of RORC in immune system function and autoimmune diseases. Order yours today and take your research to the next level.
Product Name:
Mouse Nuclear receptor ROR-gamma (Rorc) ELISA Kit
SKU:
MOEB0813
Size:
96T
Target:
Mouse Nuclear receptor ROR-gamma (Rorc)
Synonyms:
Nuclear receptor RZR-gamma, Nuclear receptor subfamily 1 group F member 3, RAR-related orphan receptor C, Retinoid-related orphan receptor-gamma, Thymus orphan receptor, TOR, Nr1f3, Rorg, Thor
Assay Type:
Sandwich
Detection Method:
ELISA
Reactivity:
Mouse
Detection Range:
0.156-10ng/mL
Sensitivity:
0.081ng/mL
Intra CV:
6.3%
Inter CV:
8.1%
Linearity:
Sample
1:2
1:4
1:8
1:16
Serum(N=5)
106-117%
92-100%
98-109%
97-107%
EDTA Plasma(N=5)
102-112%
106-115%
108-119%
83-96%
Heparin Plasma(N=5)
90-100%
104-115%
100-110%
95-107%
Recovery:
Sample Type
Average(%)
Recovery Range(%)
Serum
105
99-111
Plasma
107
101-113
Function:
Isoform 2: Essential for thymopoiesis and the development of several secondary lymphoid tissues, including lymph nodes and Peyer's patches (PubMed:10602018, PubMed:14691482, PubMed:16148126). Required for the generation of LTi (lymphoid tissue inducer) cells. Regulates thymocyte survival through DNA-binding on ROREs of target gene promoter regions and recruitment of coactivaros via the AF-2. Also plays a key role, downstream of IL6 and TGFB and synergistically with RORA, for lineage specification of uncommitted CD4(+) T-helper (T(H)) cells into T(H)17 cells, antagonizing the T(H)1 program. Probably regulates IL17 and IL17F expression on T(H) by binding to the essential enhancer conserved non-coding sequence 2 (CNS2) in the IL17-IL17F locus (PubMed:16990136, PubMed:18164222, PubMed:26607793). May also play a role in the pre-TCR activation cascade leading to the maturation of alpha/beta T-cells and may participate in the regulation of DNA accessibility in the TCR-J(alpha) locus (PubMed:9881970, PubMed:10602018, PubMed:14691482, PubMed:16148126, PubMed:16990136, PubMed:18164222). Plays an indispensable role in the induction of IFN-gamma dependent anti-mycobacterial systemic immunity.
Uniprot:
P51450
Sample Type:
Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:
Natural and recombinant mouse Nuclear receptor ROR-gamma
Sub Unit:
Interacts (via AF-2 motif) with the coactivators NCOA1, NCOA2 and PPARGC1A (via LXXLL motif). Interacts with the corepressor NCOR1. Interacts with CRY1. Interacts (via AF-2 motif) with PROX1. Interacts with FOXP3.
Research Area:
Epigenetics
Subcellular Location:
Nucleus
Storage:
Please see kit components below for exact storage details
Note:
For research use only
UniProt Protein Function:
Nuclear receptor that binds DNA as a monomer to ROR response elements (RORE) containing a single core motif half-site 5'-AGGTCA-3' preceded by a short A-T-rich sequence. Key regulator of cellular differentiation, immunity, peripheral circadian rhythm as well as lipid, steroid, xenobiotics and glucose metabolism. Considered to have intrinsic transcriptional activity, have some natural ligands like oxysterols that act as agonists (25-hydroxycholesterol) or inverse agonists (7-oxygenated sterols), enhancing or repressing the transcriptional activity, respectively. Recruits distinct combinations of cofactors to target gene regulatory regions to modulate their transcriptional expression, depending on the tissue, time and promoter contexts (PubMed:17666523, PubMed:19381306, PubMed:19965867, PubMed:21853531, PubMed:22789990, PubMed:23723244). Regulates the circadian expression of clock genes such as CRY1, ARNTL/BMAL1 and NR1D1 in peripheral tissues and in a tissue-selective manner (PubMed:22753030). Competes with NR1D1 for binding to their shared DNA response element on some clock genes such as ARNTL/BMAL1, CRY1 and NR1D1 itself, resulting in NR1D1-mediated repression or RORC-mediated activation of the expression, leading to the circadian pattern of clock genes expression. Therefore influences the period length and stability of the clock (PubMed:22753030). Involved in the regulation of the rhythmic expression of genes involved in glucose and lipid metabolism, including PLIN2 and AVPR1A. Negative regulator of adipocyte differentiation through the regulation of early phase genes expression, such as MMP3. Controls adipogenesis as well as adipocyte size and modulates insulin sensitivity in obesity. In liver, has specific and redundant functions with RORA as positive or negative modulator of expression of genes encoding phase I and Phase II proteins involved in the metabolism of lipids, steroids and xenobiotics, such as SULT1E1 (PubMed:21853531). Also plays also a role in the regulation of hepatocyte glucose metabolism through the regulation of G6PC and PCK1. Regulates the rhythmic expression of PROX1 and promotes its nuclear localization.
Nuclear receptor RZR-gamma; Nuclear receptor subfamily 1 group F member 3; RAR-related orphan receptor C; Retinoid-related orphan receptor-gamma; Thymus orphan receptor; TOR
Protein Family:
Nuclear receptor
UniProt Gene Name:
Rorc
UniProt Entry Name:
Component
Quantity (96 Assays)
Storage
ELISA Microplate (Dismountable)
8×12 strips
-20°C
Lyophilized Standard
2
-20°C
Sample Diluent
20ml
-20°C
Assay Diluent A
10mL
-20°C
Assay Diluent B
10mL
-20°C
Detection Reagent A
120µL
-20°C
Detection Reagent B
120µL
-20°C
Wash Buffer
30mL
4°C
Substrate
10mL
4°C
Stop Solution
10mL
4°C
Plate Sealer
5
-
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
*Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37°C directly). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at -20°C until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their experiments. We recommend running all samples in duplicate.
Step
1.
Add Sample: Add 100µL of Standard, Blank, or Sample per well. The blank well is added with Sample diluent. Solutions are added to the bottom of micro ELISA plate well, avoid inside wall touching and foaming as possible. Mix it gently. Cover the plate with sealer we provided. Incubate for 120 minutes at 37°C.
2.
Remove the liquid from each well, don't wash. Add 100µL of Detection Reagent A working solution to each well. Cover with the Plate sealer. Gently tap the plate to ensure thorough mixing. Incubate for 1 hour at 37°C. Note: if Detection Reagent A appears cloudy warm to room temperature until solution is uniform.
3.
Aspirate each well and wash, repeating the process three times. Wash by filling each well with Wash Buffer (approximately 400µL) (a squirt bottle, multi-channel pipette,manifold dispenser or automated washer are needed). Complete removal of liquid at each step is essential. After the last wash, completely remove remaining Wash Buffer by aspirating or decanting. Invert the plate and pat it against thick clean absorbent paper.
4.
Add 100µL of Detection Reagent B working solution to each well. Cover with the Plate sealer. Incubate for 60 minutes at 37°C.
5.
Repeat the wash process for five times as conducted in step 3.
6.
Add 90µL of Substrate Solution to each well. Cover with a new Plate sealer and incubate for 10-20 minutes at 37°C. Protect the plate from light. The reaction time can be shortened or extended according to the actual color change, but this should not exceed more than 30 minutes. When apparent gradient appears in standard wells, user should terminatethe reaction.
7.
Add 50µL of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing.
8.
Determine the optical density (OD value) of each well at once, using a micro-plate reader set to 450 nm. User should open the micro-plate reader in advance, preheat the instrument, and set the testing parameters.
9.
After experiment, store all reagents according to the specified storage temperature respectively until their expiry.
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 Type
Protocol
Serum
If 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.
Plasma
Collect 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 Fluid
Collect 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 supernatant
Collect 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 lysates
Solubilize 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 homogenates
The 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 lysates
Rinse 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 Milk
Collect 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.
