Nori Human PCNA ELISA Kit
$461.00 – $832.00
This ELISA kit is for quantification of PCNA in human. This is a quick ELISA assay that reduces time to 50% compared to the conventional method, and the entire assay only takes 3 hours. This assay employs the quantitative sandwich enzyme immunoassay technique and uses biotin-streptavidin chemistry to improve the performance of the assays. An antibody specific for PCNA has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any PCNA present is bound by the immobilized antibody. After washing away any unbound substances, a detection antibody specific for PCNA is added to the wells. Following wash to remove any unbound antibody reagent, a detection reagent is added. After intensive wash a substrate solution is added to the wells and color develops in proportion to the amount of PCNA bound in the initial step. The color development is stopped, and the intensity of the color is measured.
Alternative names for PCNA: Proliferating cell nuclear antigen
This product is for Laboratory Research Use Only not for diagnostic and therapeutic purposes or any other purposes.
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Description
Nori Human PCNAĀ ELISA Kit Summary
Alternative names for PCNA: Proliferating cell nuclear antigen
| Assay Type | Solid Phase Sandwich ELISA |
| Format | 96-well Microplate or 96-Well Strip Microplate |
| Method of Detection | Colorimetric |
| Number of Targets Detected | 1 |
| Target Antiben Accession Number | P12004 |
| Assay Length | 3 hours |
| Quantitative/Semiquantitative | Quantitative |
| Sample Type | Plasma, Serum, Cell Culture, Urine, Cell/Tissue Lysates, Synovial Fluid, BAL, |
| Recommended Sample Dilution (Plasma/Serum) | No dilution for sample <ULOQ; sufficient dilution for samples >ULOQ |
| Sensitivity | 250 pg/mL |
| Detection Range | 1.25-80 ng/mL |
| Specificity | Human PCNA |
| Cross-Reactivity | < 0.5% cross-reactivity observed with available related molecules, < 50% cross-species reactivity observed with species tested. |
| Interference | No significant interference observed with available related molecules |
| Storage/Stability | 4 ĀŗC for up to 6 months |
| Usage | For Laboratory Research Use Only. Not for diagnostic or therapeutic use. |
| Additional Notes | The kit allows for use in multiple experiments. |
Standard Curve
Kit Components
1. Pre-coated 96-well Microplate
2. Biotinylated Detection Antibody
3. Streptavidin-HRP Conjugate
4. Lyophilized Standards
5. TMB One-Step Substrate
6. Stop Solution
7. 20 x PBS
8. Assay Buffer
Other Materials Required but not Provided:
1. Microplate Reader capable of measuring absorption at 450 nm
2. Log-log graph paper or computer and software for ELISA data analysis
3. Precision pipettes (1-1000 Āµl)
4. Multi-channel pipettes (300 Āµl)
5. Distilled or deionized water
Protocol Outline
1. Prepare all reagents, samples and standards as instructed in the datasheet.
2. Add 100 Āµl of Standard or samples to each well and incubate 1 h at RT.
3. Add 100 Āµl of Working Detection Antibody to each well and incubate 1 h at RT.
4. Add 100 Āµl of Working Streptavidin-HRP to each well and incubate 20 min at RT.
5. Add 100 Āµl of Substrate to each well and incubate 5-30 min at RT.
6. Add 50 Āµl of Stop Solution to each well and read at 450 nm immediately.
Background:Ā
Proliferating cell nuclear antigenĀ (PCNA) is aĀ DNA clampĀ that acts as aĀ processivityĀ factor forĀ DNA polymerase Ī“Ā inĀ eukaryoticĀ cellsĀ and is essential for replication. PCNA is aĀ homotrimerĀ and achieves its processivity by encircling the DNA, where it acts as a scaffold to recruit proteins involved in DNA replication, DNA repair, chromatin remodeling andĀ epigenetics.[1] PCNA is a cofactor of DNA polymerase delta and acts as a homotrimer and helps increase the processivity of leading strand synthesis during DNA replication. In response to DNA damage, this protein isĀ ubiquitinatedĀ and is involved in the RAD6-dependent DNA repair pathway. PCNA was originally identified as anĀ antigenĀ that is expressed in theĀ nucleiĀ of cells during theĀ DNA synthesis phaseĀ of theĀ cell cycle.[2]Ā Ā PCNA helps holdĀ DNA polymeraseĀ epsilon (Pol Īµ) to DNA. PCNA is clamped[3]Ā toĀ DNAĀ through the action ofĀ replication factor C(RFC),[4]Ā which is a heteropentameric member of theĀ AAA+Ā class of ATPases. Expression of PCNA is under the control ofĀ E2FĀ transcription factor-containing complexes.[5] SinceĀ DNA polymeraseĀ epsilon is involved in resynthesis of excised damaged DNA strands duringĀ DNA repair, PCNA is important for both DNA synthesis and DNA repair.[6][7] Many proteins interact with PCNA via the two known PCNA-interacting motifs PCNA-interacting peptide (PIP) boxĀ and AlkB homologue 2 PCNA interacting motif (APIM).[8]Ā Proteins binding to PCNA via the PIP-box are mainly involved in DNA replication whereas proteins binding to PCNA via APIM are mainly important in the context of genotoxic stress.[9]
References
- Moldovan GL, Pfander B, Jentsch S (2007).Ā Cell.Ā 129Ā (4): 665ā79.
- Leonardi E, et al. (1992).Ā J. Clin. Pathol.Ā 45Ā (5): 416ā419.
- Bowman GD, O’Donnell M, Kuriyan J (2004). Nature.Ā 429Ā (6993): 724ā730.
- Zhang G, et al. (1999).Ā Proc. Natl. Acad. Sci. U.S.A.Ā 96Ā (5): 1869ā1874.
- Egelkrout EM, et al. (2002).Ā Ā Plant Cell.Ā 14Ā (12): 3225ā3236.
- Shivji KK, Kenny MK, Wood RD (1992). Cell.Ā 69Ā (2): 367ā74.
- Essers J, et al. (2005).Ā Mol. Cell. Biol.Ā 25Ā (21): 9350ā9359.
- Gilljam KM, et al. (2009) The Journal of Cell Biology.Ā 186Ā (5): 645ā54.
- Mailand N, et al. (2013). Nature Reviews. Molecular Cell Biology.Ā 14Ā (5): 269ā82.
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