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Rat DNA topoisomerase 2-alpha (Top2a) ELISA Kit (RTEB1585)

SKU:
RTEB1585
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P41516
ELISA Type:
Sandwich
Reactivity:
Rat
€649

Description

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Rat DNA topoisomerase 2-alpha (Top2a) ELISA Kit

The Rat DNA Topoisomerase 2 Alpha (TOP2A) ELISA Kit is a highly sensitive and specific tool for the detection of TOP2A levels in rat serum, plasma, and cell culture supernatants. This kit is designed to provide accurate and reproducible results, making it an essential resource for a variety of research applications.TOP2A is an important enzyme involved in DNA replication and repair, playing a crucial role in maintaining genomic integrity. Dysregulation of TOP2A has been linked to a variety of diseases, including cancer and neurodegenerative disorders, making it a valuable biomarker for studying these conditions and exploring potential therapies.

With its high sensitivity and specificity, the Rat DNA Topoisomerase 2 Alpha (TOP2A) ELISA Kit offers researchers a reliable and efficient method for quantifying TOP2A levels, facilitating in-depth investigations into the role of this enzyme in health and disease.

Product Name:Rat DNA topoisomerase 2-alpha (Top2a) ELISA Kit
SKU:RTEB1585
Size:96T
Target:Rat DNA topoisomerase 2-alpha (Top2a)
Synonyms:DNA topoisomerase II, alpha isozyme, Top-2, Top2
Assay Type:Sandwich
Detection Method:ELISA
Reactivity:Rat
Intra CV:3.9%
Inter CV:7.4%
Linearity:
Sample1:21:41:81:16
Serum(N=5)116-126%89-101%84-93%108-116%
EDTA Plasma(N=5)94-103%91-103%92-104%88-97%
Heparin Plasma(N=5)87-97%106-118%100-112%97-106%
Recovery:
Sample TypeAverage(%)Recovery Range(%)
Serum9488-100
Plasma9690-102
Function:Control of topological states of DNA by transient breakage and subsequent rejoining of DNA strands. Topoisomerase II makes double-strand breaks. Essential during mitosis and meiosis for proper segregation of daughter chromosomes. May play a role in regulating the period length of ARNTL/BMAL1 transcriptional oscillation.
Uniprot:P41516
Sample Type:Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:Natural and recombinant rat DNA topoisomerase 2-alpha
Sub Unit:Homodimer. Interacts with COPS5. Interacts with RECQL5; this stimulates DNA decatenation. Interacts with SETMAR; stimulates the topoisomerase activity.
Research Area:Cancer
Subcellular Location:Nucleus
Storage:Please see kit components below for exact storage details
Note:For research use only
UniProt Protein Function:Control of topological states of DNA by transient breakage and subsequent rejoining of DNA strands. Topoisomerase II makes double-strand breaks. Essential during mitosis and meiosis for proper segregation of daughter chromosomes. May play a role in regulating the period length of ARNTL/BMAL1 transcriptional oscillation.
UniProt Protein Details:
NCBI Summary:DNA topoisomerase, a nuclear enzyme that can alter the topologic state of DNA during transcription [RGD, Feb 2006]
UniProt Code:P41516
NCBI GenInfo Identifier:38259192
NCBI Gene ID:360243
NCBI Accession:NP_071519.2
UniProt Secondary Accession:P41516
UniProt Related Accession:P41516
Molecular Weight:173,221 Da
NCBI Full Name:DNA topoisomerase 2-alpha
NCBI Synonym Full Names:DNA topoisomerase II alpha
NCBI Official Symbol:Top2a
NCBI Official Synonym Symbols:
NCBI Protein Information:DNA topoisomerase 2-alpha
UniProt Protein Name:DNA topoisomerase 2-alpha
UniProt Synonym Protein Names:DNA topoisomerase II, alpha isozyme
Protein Family:DNA topoisomerase
UniProt Gene Name:Top2a
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.

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