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Porcine Cell division control protein 42 homolog (CDC42) ELISA Kit (PREB0098)

SKU:
PREB0098
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
Q007T2
ELISA Type:
Sandwich
Reactivity:
Porcine
€699
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Description

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Porcine Cell division control protein 42 homolog (CDC42) ELISA Kit

The Porcine Cell Division Control Protein 42 Homolog (CDC42) ELISA Kit is specifically designed for the accurate quantification of CDC42 levels in porcine samples. This kit offers high sensitivity and specificity, ensuring precise and reliable results for a variety of research applications. CDC42 is a key regulator of cell division and plays a crucial role in various cellular processes, including cell migration, proliferation, and polarity.

Dysregulation of CDC42 has been implicated in numerous diseases, making it a valuable biomarker for studying and potentially targeting these conditions. With the Porcine Cell Division Control Protein 42 Homolog (CDC42) ELISA Kit, researchers can confidently assess CDC42 levels in porcine samples, furthering our understanding of cellular signaling pathways and potentially identifying novel therapeutic targets.

Product Name:Porcine Cell division control protein 42 homolog (CDC42) ELISA Kit
SKU:PREB0098
Size:96T
Target:Porcine Cell division control protein 42 homolog (CDC42)
Synonyms:Cell division control protein 42 homolog, CDC42
Detection Method:ELISA
Reactivity:Pig
Intra CV:Provided with the Kit
Inter CV:Provided with the Kit
Linearity:Provided with the Kit
Recovery:Provided with the Kit
Function:Plasma membrane-associated small GTPase which cycles between an active GTP-bound and an inactive GDP-bound state. In active state binds to a variety of effector proteins to regulate cellular responses. Involved in epithelial cell polarization processes. Regulates the bipolar attachment of spindle microtubules to kinetochores before chromosome congression in metaphase. Plays a role in the extension and maintenance of the formation of thin, actin-rich surface projections called filopodia. Mediates CDC42-dependent cell migration (By similarity). Required for DOCK10-mediated spine formation in Purkinje cells and hippocampal neurons. Facilitates filopodia formation upon DOCK11-activation.
Uniprot:Q007T2
Sample Type:Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:Natural and recombinant pig Cell division control protein 42 homolog
Sub Unit:Interacts with CDC42EP1, CDC42EP2, CDC42EP3, CDC42EP4, CDC42EP5, CDC42SE1, CDC42SE2, PARD6A, PARD6B and PARD6G (in a GTP-dependent manner). Interacts with activated CSPG4 and with BAIAP2. Interacts with Zizimin1/DOCK9 and Zizimin2/DOCK11, which activate it by exchanging GDP for GTP. Interacts with NET1 and ARHGAP33/TCGAP. Part of a complex with PARD3, PARD6A or PARD6B and PRKCI or PRKCZ. The GTP-bound form interacts with CCPG1. Interacts with USP6 (By similarity). Interacts with NEK6 (By similarity). Part of a collagen stimulated complex involved in cell migration composed of CDC42, CRK, TNK2 and BCAR1/p130cas. Interacts with ITGB1BP1 (By similarity). Interacts with ARHGDIA; this interaction inactivates and stabilizes CDC42. Interacts with ARHGDIB; this maintains CDC42 in the inactive, GDP-bound form (By similarity). Interacts in (GTP-bound form) with FNBP1L and ABI1, but only in the presence of FNBP1L.
Research Area:Cancer
Subcellular Location:Cell membrane Lipid-anchor Cytoplasmic side Cytoplasm Cytoskeleton Microtubule organizing center Centrosome Midbody Localizes to spindle during prometaphase cells. Moves to the central spindle as cells progressed through anaphase to telophase. Localizes at the end of cytokinesis in the intercellular bridge formed between two daughter cells. Its localization is regulated by the activities of guanine nucleotide exchange factor ECT2 and GTPase activating protein RACGAP1. Colocalizes with NEK6 in the centrosome (By similarity).
Storage:Please see kit components below for exact storage details
Note:For research use only
UniProt Protein Function:Function: Plasma membrane-associated small GTPase which cycles between an active GTP-bound and an inactive GDP-bound state. In active state binds to a variety of effector proteins to regulate cellular responses. Involved in epithelial cell polarization processes. Regulates the bipolar attachment of spindle microtubules to kinetochores before chromosome congression in metaphase. Plays a role in the extension and maintenance of the formation of thin, actin-rich surface projections called filopodia. Mediates CDC42-dependent cell migration Enzyme regulation: Regulated by guanine nucleotide exchange factors (GEFs) which promote the exchange of bound GDP for free GTP, GTPase activating proteins (GAPs) which increase the GTP hydrolysis activity, and GDP dissociation inhibitors which inhibit the dissociation of the nucleotide from the GTPase
UniProt Protein Details:
NCBI Summary:
UniProt Code:Q007T2
NCBI GenInfo Identifier:118403806
NCBI Gene ID:780428
NCBI Accession:NP_001072148.1
UniProt Secondary Accession:Q007T2,D1YSL6
UniProt Related Accession:Q007T2
Molecular Weight:21,327 Da
NCBI Full Name:cell division control protein 42 homolog
NCBI Synonym Full Names:
NCBI Official Symbol:CDC42
NCBI Official Synonym Symbols:
NCBI Protein Information:cell division control protein 42 homolog
UniProt Protein Name:Cell division control protein 42 homolog
UniProt Synonym Protein Names:
Protein Family:Cdc42
UniProt Gene Name:CDC42
UniProt Entry Name:CDC42_PIG
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.

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