The Rat DLG4 (Discs Large Homolog 4) ELISA Kit is a reliable tool for the accurate detection of DLG4 levels in rat tissue samples. This kit offers high sensitivity and specificity, allowing for robust and reproducible results in a variety of research applications.DLG4, also known as PSD95, is a key protein involved in synaptic function and plasticity in the brain. It plays a critical role in regulating neurotransmitter release and synaptic strength, making it essential for studying conditions such as Alzheimer's disease, schizophrenia, and autism spectrum disorders.
By using the Rat DLG4 ELISA Kit, researchers can gain valuable insights into the role of DLG4 in neurological disorders and potentially develop new therapeutic strategies targeting this protein. With its user-friendly protocol and reliable performance, this kit is an essential tool for any laboratory studying synaptic function and brain disorders in rats.
Product Name:
Rat Disks large homolog 4 (Dlg4) ELISA Kit
SKU:
RTEB0794
Size:
96T
Target:
Rat Disks large homolog 4 (Dlg4)
Synonyms:
Postsynaptic density protein 95, Synapse-associated protein 90, PSD-95, SAP-90, Dlgh4, Psd95
Assay Type:
Sandwich
Detection Method:
ELISA
Reactivity:
Rat
Detection Range:
0.156-10ng/mL
Sensitivity:
0.052ng/mL
Intra CV:
3.9%
Inter CV:
7.2%
Linearity:
Sample
1:2
1:4
1:8
1:16
Serum(N=5)
99-109%
93-104%
80-93%
96-106%
EDTA Plasma(N=5)
118-128%
110-118%
96-107%
108-117%
Heparin Plasma(N=5)
87-99%
101-113%
88-100%
89-99%
Recovery:
Sample Type
Average(%)
Recovery Range(%)
Serum
99
93-105
Plasma
101
95-107
Function:
Interacts with the cytoplasmic tail of NMDA receptor subunits and shaker-type potassium channels. Required for synaptic plasticity associated with NMDA receptor signaling. Overexpression or depletion of DLG4 changes the ratio of excitatory to inhibitory synapses in hippocampal neurons. May reduce the amplitude of ASIC3 acid-evoked currents by retaining the channel intracellularly. May regulate the intracellular trafficking of ADR1B.
Uniprot:
P31016
Sample Type:
Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:
Natural and recombinant rat Disks large homolog 4
Sub Unit:
Interacts through its PDZ domains with ANO2 and NETO1. Interacts with KCNJ4 (PubMed:11997254). Interacts through its first two PDZ domains with GRIN2A, GRIN2B, GRIN2C, GRIN2D, ASIC3, certain splice forms of GRIN1, KCND2, CXADR, SYNGAP1, KCNA1, KCNA2, KCNA3, KCNA4, ERBB4, LRRC4; LRRC4B and SEMA4C. Interacts through its first PDZ domain with GRIK2, KCNA4 and CRIPT. Interacts through its second PDZ domain with the PDZ domain of NOS1 or the C-terminus of CAPON. Interacts through its third PDZ domain with NLGN1 and CRIPT, and probably with NLGN2 and NLGN3. Interacts through its guanylate kinase-like domain with DLGAP1/GKAP, DLGAP2, DLGAP3, DLGAP4, MAP1A, BEGAIN, SIPA1L1 and KIF13B (PubMed:9115257, PubMed:9756850). Isoform 2 interacts through an L27 domain with HGS/HRS and the first L27 domain of CASK. Interacts with ADR1B, ANKS1B and PRR7 (PubMed:15629447). May interact with HTR2A. Interacts with ADAM22, KLHL17 and LGI1. Interacts with FRMPD4 (via C-terminus) (PubMed:19118189). Interacts with LRFN1, LRFN2 and LRFN4, but not with LRFN3 nor LRFN5. Interacts (via N-terminal tandem pair of PDZ domains) with GPER1 (via C-terminus tail motif); the interaction is direct and induces the increase of GPER1 protein levels residing at the plasma membrane surface in a estradiol-independent manner (PubMed:23300088). Interacts (via N-terminus tandem pair of PDZ domains) with NOS1 (via N-terminal domain) (PubMed:23300088). Interacts with SHANK3. Interacts with GPR85 (By similarity). Interacts with CACNG2 and MPP2 (via the SH3-Guanylate kinase-like sub-module) (PubMed:27756895).
Research Area:
Neurosciences
Subcellular Location:
Cell membrane Peripheral membrane protein Cell junction Synapse Postsynaptic cell membrane Postsynaptic density Cell junction Synapse Membrane Cell projection Axon High levels in postsynaptic density of neurons in the forebrain. Also in presynaptic region of inhibitory synapses formed by cerebellar basket cells on axon hillocks of Purkinje cells.
Storage:
Please see kit components below for exact storage details
Note:
For research use only
UniProt Protein Function:
Function: Interacts with the cytoplasmic tail of NMDA receptor subunits and shaker-type potassium channels. Required for synaptic plasticity associated with NMDA receptor signaling. Overexpression or depletion of DLG4 changes the ratio of excitatory to inhibitory synapses in hippocampal neurons. May reduce the amplitude of ASIC3 acid-evoked currents by retaining the channel intracellularly. May regulate the intracellular trafficking of ADR1B. Ref.17 Ref.19 Ref.27Subunit structure: Interacts through its PDZ domains with ANO2 and NETO1. Interacts through its first two PDZ domains with GRIN2A, GRIN2B, GRIN2C, GRIN2D, ASIC3, certain splice forms of GRIN1, KCND2, CXADR, SYNGAP1, KCNA1, KCNA2, KCNA3, KCNA4, ERBB4, LRRC4; LRRC4B and SEMA4C. Interacts through its first PDZ domain with GRIK2, KCNA4 and CRIPT. Interacts through its second PDZ domain with the PDZ domain of NOS1 or the C-terminus of CAPON. Interacts through its third PDZ domain with NLGN1 and CRIPT, and probably with NLGN2 and NLGN3. Interacts through its guanylate kinase-like domain with DLGAP1/GKAP, DLGAP2, DLGAP3, DLGAP4, MAP1A, BEGAIN, SIPA1L1 and KIF13B. Isoform 2 interacts through an L27 domain with HGS/HRS and the first L27 domain of CASK. Interacts with ADR1B, ANKS1B and PRR7. May interact with HTR2A. Interacts with ADAM22, KLHL17 and LGI1. Interacts with FRMPD4 (via C-terminus). Interacts with LRFN1, LRFN2 and LRFN4, but not with LRFN3 nor LRFN5. Interacts (via N-terminus tandem pair of PDZ domains) with GPER1 (via C-terminus tail motif); the interaction is direct and induces the increase of GPER1 protein levels residing at the plasma membrane surface in a estradiol-independent manner. Interacts (via N-terminus tandem pair of PDZ domains) with NOS1 (via N-terminus domain). Interacts with SHANK3.Subcellular location: Cell membrane; Peripheral membrane protein. Cell junction › synapse › postsynaptic cell membrane › postsynaptic density. Cell junction › synapse. Note: High levels in postsynaptic density of neurons in the forebrain. Also in presynaptic region of inhibitory synapses formed by cerebellar basket cells on axon hillocks of Purkinje cells. Ref.6 Ref.7 Ref.14Tissue specificity: Detected in brain (at protein level). Brain. Highest levels of isoform 2 in cerebellum, cortex, hippocampus, and corpus striatum. Ref.14 Ref.16Domain: The PDZ domain 3 mediates interaction with ADR1B By similarity.The L27 domain near the N-terminus of isoform 2 is required for HGS/HRS-dependent targeting to postsynaptic density By similarity.Post-translational modification: Palmitoylation of isoform 1 is required for targeting to postsynaptic density. Ref.12Sequence similarities: Belongs to the MAGUK family.Contains 1 guanylate kinase-like domain.Contains 3 PDZ (DHR) domains.Contains 1 SH3 domain.
UniProt Protein Details:
NCBI Summary:
a molecular scaffolding protein that binds and clusters N-methyl-D-aspartate receptors at neuronal synapses; may be involved in guanine nucleotide-mediated signal transduction pathway [RGD, Feb 2006]
disks large homolog 4; PSD-95; SAP-90; discs large homolog 4; synapse-associated protein 90; postsynaptic density protein 95; synapse-associated protein SAP90
UniProt Protein Name:
Disks large homolog 4
UniProt Synonym Protein Names:
Postsynaptic density protein 95; PSD-95; Synapse-associated protein 90
Protein Family:
Disks large
UniProt Gene Name:
Dlg4
UniProt Entry Name:
DLG4_RAT
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.
