科技澳门威斯尼人 > PCR芯片实验澳门威斯尼人 > 炎症反应和自身免疫ChIP 澳门威斯尼人 Inflammatory Response & Autoimmunity EpiTect ChIP qPCR Array

炎症反应和自身免疫ChIP 澳门威斯尼人 Inflammatory Response & Autoimmunity EpiTect ChIP qPCR Array

炎症反应和自身免疫ChIP 澳门威斯尼人 Inflammatory Response & Autoimmunity EpiTect ChIP qPCR Array
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简介:Inflammatory Response & Autoimmunity EpiTect ChIP qPCR Array 炎症反应和自身免疫ChIP 澳门威斯尼人
提供商:SAbiosciences
澳门威斯尼人名称:炎症反应和自身免疫ChIP 澳门威斯尼人
地区:美国
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Inflammatory Response & Autoimmunity EpiTect ChIP qPCR Array

炎症反应和自身免疫ChIP 澳门威斯尼人
 
ProductSpeciesTechnologyCat. No.
Inflammatory Response & Autoimmunity EpiTect ChIP qPCR ArrayHumanHistone ModificationsGH-504A
Inflammatory Response & Autoimmunity EpiTect ChIP qPCR ArrayMouseHistone ModificationsGM-504A
The Human Response and Autoimmunity EpiTect Chip qPCR Array profiles modified histone and nuclear protein binding to the proximal promoter of a panel of genes involved in inflammation and Inflammatory Response and Autoimmunitys. The array contains 96 pairs of qPCR primers targeting the 1-kb region downstream of the transcription start sites (TSS) of 84 biological important genes plus 12 appropriate control regions. The genes represented by this array demonstrate at least differential regulation at the transcriptional level if not also known associations with modified histones. This array includes cytokine, cytokine receptor, transcription factor, and other signaling molecule genes mediating immune cascade reactions during inflammation and Inflammatory Response and Autoimmunitys. Thoroughly researched panels of genes involved in the acute-phase, inflammatory, and humoral immune responses are represented as well. Profiling the histone modifications and nuclear protein binding events at these gene promoters in your immune cell populations will help you correlate these interactions with each gene's known inflammatory role. The results also provide insights into the epigenetic molecular mechanisms and biological pathways behind inflammatory Response and Autoimmunity, especially chronic inflammation. Using Chromatin Immunoprecipitation and this real-time PCR Array, you can easily and reliably analyze the association between your chosen nuclear factors and the promoters for a focused gene panel involved in inflammation.
炎症反应和自身免疫ChIP 澳门威斯尼人用于研究参与炎症和炎性反应及自身免疫
中修饰组蛋白和核酸蛋白绑定到近端启动子的基因。该芯片包含96对qPCR引物,这些引物针对84个生物学重要基因的转录起始站点(TSS) 下游1 kb区域,以及12个对照域。如果不知道修饰后的组蛋白,这个芯片至少也能证明转录水平的不同调控。这个芯片包括细胞因子、细胞因子受体、转录因子及在炎症和炎性反应和自身免疫介导免疫级联反应的其他信号分子,彻底研究急性期、炎症以及体液免疫反应的基因。分析你的免疫细胞群中组蛋白修饰和核蛋白绑定这些基因启动子,有助于分析这些炎症作用已知的基因的相互作用。分析结果还助于洞察炎症反应和自身免疫,尤其是慢性炎症背后的表观遗传分子机制和生物学通路。通过染色质免疫沉淀和EpiTect ChIP 澳门威斯尼人,可以很简易、可靠地分析你选择的核因子和炎症关键基因启动子的关联。
 
EpiTect ChIP qPCR仅用于分子生物学应用。本产品不用于疾病的诊断、预防和治疗。
 
Chemokines: C5, CCL1, CCL11, CCL17, CCL19, CCL2, CCL20, CCL21, CCL3, CCL3, CCL4, CCL5, CXCL1, CXCL10, CXCL11, CXCL12, CXCL13, CXCL14, CXCL5, CXCL6, CXCL9, IL13, IL8.
Chemokine Receptors: CCR3, CCR4, CCR5, CCR6, CCR9, CX3CR1, IL10RB, IL1R1, IL1RN, IL6R, IL8RB.
Cytokine Genes: CD40LG, CSF2, FASLG, IFNA2, IFNG, IL10, IL13, IL15, IL17A, IL17C, IL17F, IL18, IL1A, IL1B, IL1F10, IL1RN, IL2, IL21, IL22, IL3, IL4, IL5, IL6, IL8, IL9, LTA, LTB, MIF, SPP1, TGFB1, TNF.
Cytokine Receptors: CCR3, CCR4, CCR5, CCR6, CXCR4, IFNA2, IL10RA, IL10RB, IL13, IL13RA1, IL5RA, IL8RB, IL9, TNFRSF1B.
Transcription Factors: FOXP3, GATA3, JUN, RELA.
Acute-Phase Response: CEBPB, CRP, IL22, IL6.
Inflammatory Response and Autoimmunity: ABCF1, BCL6, C3, CCL11, CCL17, CCL19, CCL2, CCL21, CCL3, CCL4, CCL5, CCR3, CCR4, CD40, CD40LG, CEBPB, CRP, CXCL1, CXCL10, CXCL5, CXCL6, CXCL9, FOS, IL10, IL10RB, IL18RAP, IL1A, IL1B, IL1F10, IL1R1, IL1RN, IL22, IL8, IL8RB, IL9, LTB4R, NFKB1, PTGS2, TLR4, TMPRSS11D, TNF, TOLLIP.
Humoral Immune Response: C3, CCL2, CCL3, CD40, IL10, IL18, IL1B, IL6, NFKB1
 

How it Works

The ChIP PCR array is a set of optimized real-time PCR primer assays on 96-well or 384-well plates for pathway or disease focused analysis of in vivo protein-DNA interactions. The ChIP PCR array performs ChIP DNA analysis with real-time PCR sensitivity and the multi-genomic loci profiling capability of a ChIP-on-chip. Simply mix your ChIP DNA samples with the appropriate ready-to-use PCR master mix, aliquot equal volumes to each well of the same plate, and then run the real-time PCR cycling program. (Download user manual)

What ChIP PCR Array Offers?

  • Function or Disease Focused: Arrays represent a panel of genomic regions relevant to a biological function or disease state.

  • Reliable & Sensitive: Arrays can analyze multiple genomic regions simultaneously with Real-Time PCR precision and sensitivity.

  • Easy to Use Data Analysis: Download an easy-to-use Excel-based data analysis template [here]. Data analysis is based on the ΔΔCt method with normalization of the specific antibody and control IgG raw data to input raw data.

Layout and Controls: The PCR Arrays are available in both 96- and 384-well plates and are used to monitor the expression of 84 genes related to a disease state or pathway plus five housekeeping genes. Controls are also included on each array for ChIP DNA quality controls and general PCR performance.
You can easily perform a ChIP PCR Array experiment in your own laboratory, or send your samples to us and take advantage of our 
PCR Array Services.

 

Performance
EpiTect Chip qPCR Arrays provide the high sensitivity, specificity and reproducibility using SYBR-based real-time PCR technology.

Sensitivity
Together with our easy and fast One-Day ChIP kit, ChIP-Grade Antibody Kits, one million cells per assay as starting material provides 100% effective call rates.

Ct Range

 

Percent Distribution of Ct Values
InputH3K4me3Control IgG
<240%27%0%
25-30100%60%0%
30-350%13%96%
Absent Calls0%0%4%

Table 1. ChIP PCR Arrays Analyze the Enrichment of 84 Genomic Sites with as Little as One Million Cells. P19 mouse embryonic carcinoma cells were prepared for ChIP Assay using the EpiTect Chip One-Day Kit and anti-H3K4me3 Antibody Kit. One million cells were used as starting material for each ChIP Assay. The purified ChIP DNA samples were characterized using Mouse Stem Cell Transcription Factor ChIP PCR Array with 1/100th of the ChIP DNA as template in each well. The Real-Time PCR results demonstrate 100 % effective call rates for the Input Fraction (Ct < 30). The difference of Ct value between the anti-H3K4me3 antibody and the control IgG fractions indicates the specific enrichment of the antibody, whereas the high Ct value of the control IgG fraction indicates the low background of the assay.

Reproducibility
The complete ChIP PCR Array System demonstrates a high degree of reproducibility across technical replicates, lots, instruments, and different handling, insuring reliable detection of differences in genomic DNA enrichment among biological samples.

Figure 5. Consistent Performance within the Same Plate or across Different Plates. Sonicated chromatin from HeLa cells (20 µg) was immunoprecipitated with 2 µg of anti-H3ac antibody or control IgG for 2 hours using the EpiTect Chip One-Day Kit. The obtained ChIP DNA samples were characterized in triplicates with EpiTect Chip qPCR primers specific for the active genes (GAPDH, RPL30, ALDOA), inactive genes (MYOD1, SERPINA), repetitive sequence (SAT2, SATa), and an ORF-free region (IGX1A) either within the same array plate or among different array plates in order to evaluate the intra- and inter-plate consistency. The anti-H3ac antibody enriched genomic DNA at active gene promoter regions with a high signal-to-noise ratio and a low co-efficiency of variation (less than 2.02%), irrespective of the type of assay (intra or inter-plate)

Figure 6. Consistent Performance with Various Amount of DNA Samples, Instruments or Handling Conditions. All experiments were performed in triplicates. Cells from MCF-7 (1 million per sample) were subjected to ChIP assay with anti-RNA Polymerase II (Pol 2) antibody followed by qPCR analysis of the proximal promoter of GAPDH, and an ORF-free region (IGX1A). Researcher A & B performed the PCR assays either in 96-well plate or 384-well plate format, on a Stratagene MX 3005 or an ABI 7900 Real-Time PCR instrument respectively. The same ChIP DNA samples were used which were stored for extended periods of time as indicated. The results demonstrate high reproducibility of PCR performance across technical replicates, lots, instruments, and differential handling.

Specific and Accurate ChIP-qPCR Detection
One prerequisite for ChIP PCR Array technology is the uniform and high
PCR amplification efficiency allowing a reciprocal comparison of ChIP enrichment among all genomic loci analyzed. The unique combination of SABiosciences' proprietary ChIP-qPCR primer design algorithm, rigorous validation of every ChIP-qPCR primer assay, and high performance SYBR Green master mix guarantees superior performance of EpiTect Chip qPCR Arrays.

A:

B:

Figure 7. Uniform Amplification Efficiency and Specific PCR Detection. 96 ChIP-qPCR primers were randomly picked from our genome-wide primer pool and analyzed for their performance. (A) All assays exhibit an average amplification efficiency of 99% with a 104.5% confidence interval between 102.5-105.2%, the uniform high amplification efficiency ensures accurate analysis of multiple genomic loci simultaneously using ΔΔCt method. (B) Each ChIP-qPCR primer assay is experimentally validated using dissociation (melt) curve analysis and agarose gel verification. Each pair of primers on PCR Array produces a single specific product as indicated by a single Dissociation Curve peak at a melting temperature (Tm) greater than 75 ºC, and PCR product was further validated on agarose gel for a single product of the predicted size without secondary products such as primer dimers

Application Examples

EpiTect Chip qPCR Arrays provide streamlined approaches to 1) Study biology or disease-focused gene regulation through histone modification and transcriptional regulatory network; 2) Monitor the dynamics of chromatin structure in the screening of function-specific epigenetic patterns; 3) Validate ChIP-on-chip or ChIP-seq results. The EpiTect Chip qPCR Arrays are also powerful tools for studying the mechanism contributing to gene expression changes observed by RT² Profiler PCR Arrays.

Below are listed a few examples of application data generated by our R&D group. To see the research using ChIP PCR Arrays published by the scientific community, please see our Publication List:http://www.sabiosciences.com/support_publication.php

Stem Cell Research

Stem cell differentiation into specific tissues involves the complex yet coordinated action of many transcription factors regulating not only tissue-specific genes, but also genes essential for differentiation itself. Histone modifications at the promoters of transcription factors are key mechanism regulating their expression. We used EpiTect Chip qPCR Arrays and RT² PCR Arrays to monitor the dynamic coordination of epigenetic modification and gene expression during retinoic acid (RA) induced differentiation of P19 mouse embryonic carcinoma cells (Figure 1). This RA treatment differentiates pluripotent P19 cells into somatic cells (Figure 2). The EpiTect Chip qPCR Array data showed that both gene expression and histone modifications on key transcription factors were changed in a dynamic manner through the course of P19 cell differentiation (Figure 3).

Figure 1. Schematic Representation of Pluripotency-Associated Gene Dynamics throughout Stem Cell Differentiation

Figure 2. Retinoic Acid (RA) Differentiation of Mouse Embryonic Carcinoma P19 Cells.

Figure 3. Dynamic Epigenetic Alternations and Gene Expression Changes during RA-Induced P19 Differentiation. ChIP PCR Arrays and RT² PCR Arrays were used to monitor the changes in gene expression levels and histone modification marks (H3Ac, H3K4me3, H3K27me3, and H3K9me3). The promoter region and expression levels of 84 key stem cell transcription factors were simultaneously analyzed during RA-induced neurogenesis of P19 cells at various time points (day 0, 4, and 8). Primer sets for the +1kb region downstream of the transcription start sites of the 84 genes and 12 control regions were preloaded on the ChIP PCR Array. Cluster analysis () of histone marks and mRNA level changes for the 84 genes were visualized as a heat map to represent the fold-differences during the RA-induced differentiation at the specified time points.

Characterize the Pattern of Histone Modifications

EpiTect Chip qPCR Arrays can be used to monitoring differential histone modifications across a gene.

Figure 4. The Custom EpiTect Chip 30Kb Tiling Array Quickly Maps Histone Modifications Surrounding the Transcription Start Site (TSS) of CDKN1A Gene. EpiTect Chip Antibodies against modified histones (H3Ac, H3K4me2, H3K27me3), or NIS were used to precipitate chromatin from one million HeLa cells. Each ChIP DNA fraction was analyzed with Custom EpiTect Chip 30Kb Tiling Array representing 30 one-kb tile intervals across the promoter region of the CDKN1A gene. The results indicate the enrichment of histone markers for actively transcribed genes (H3Ac and H3K4me2) but not marks for transcriptional inactive genes (H3K27me3) in the genomic region surrounding the TSS of CDNK1A.


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