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Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Gly73 of human Helios protein.
Product Usage Information
For optimal ChIP and ChIP-seq results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits. The CUT&RUN dilution was determined using CUT&RUN Assay Kit #86652. The CUT&Tag dilution was determined using CUT&Tag Assay Kit #77552.
| Application | Dilution |
|---|---|
| Western Blotting | 1:1000 |
| Simple Western™ | 1:10 - 1:50 |
| Immunoprecipitation | 1:200 |
| Flow Cytometry (Fixed/Permeabilized) | 1:50 - 1:200 |
| Chromatin IP | 1:50 |
| Chromatin IP-seq | 1:50 |
| CUT&RUN | 1:50 |
| CUT&Tag | 1:50 |
Specificity/Sensitivity
Species Reactivity:
Human, Mouse
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
参考图片
CUT&Tag was performed with Jurkat cells and Helios (D8W4X) XP® Rabbit mAb, using CUT&Tag Assay Kit #77552. DNA library was prepared using CUT&Tag Dual Index Primers and PCR Master Mix for Illumina Systems #47415. The figures show binding across chromosome 17 (upper), including IKZF3 (lower), a known target gene of Helios (see our ChIP-qPCR figure).
Western blot analysis of extracts from various cell lines using Helios (D8W4X) XP® Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Western blot analysis of extracts from 293T cells, untransfected (-) or transfected with a construct expressing Myc-tagged full-length human Helios (hHelios-Myc; +), using Helios (D8W4X) XP® Rabbit mAb.
Simple Western™ analysis of lysates (0.1 mg/mL) from Jurkat cells using Helios (D8W4X) XP® Rabbit mAb #42427. The virtual lane view (left) shows the target (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
Immunoprecipitation of Helios from Jurkat cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Helios (D8W4X) XP® Rabbit mAb. Western blot analysis was performed using Helios (D8W4X) XP® Rabbit mAb.
Flow cytometric analysis of RL (blue) and Jurkat (green) cells using Helios (D8W4X) XP® Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary anitbody.
Chromatin immunoprecipitations were performed with cross-linked chromatin from Jurkat cells and Helios (D8W4X) XP® Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across IKZF3, a known target gene of Helios (see additional figure containing ChIP-qPCR data).
Chromatin immunoprecipitations were performed with cross-linked chromatin from Jurkat cells and Helios (D8W4X) XP® Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across chromosome 17 (upper), including IKZF3 (lower), a known target gene of Helios (see additional figure containing ChIP-qPCR data).
Chromatin immunoprecipitations were performed with cross-linked chromatin from Jurkat cells and either Helios (D8W4X) XP® Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human IKZF3 Promoter Primers #99663, human IL-2 promoter primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
CUT&RUN was performed with Jurkat cells and Helios (D8W4X) XP® Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Library was prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across IKZF3 gene.
CUT&RUN was performed with Jurkat cells and Helios (D8W4X) XP® Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Library was prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 17 (upper), including IKZF3 gene (lower).
CUT&RUN was performed with Jurkat cells and either Helios (D8W4X) XP® Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using human TMEM exon 1, human CKS1B exon 1, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent
CUT&Tag was performed with Jurkat cells and Helios (D8W4X) XP® Rabbit mAb, using CUT&Tag Assay Kit #77552. DNA library was prepared using CUT&Tag Dual Index Primers and PCR Master Mix for Illumina Systems #47415. The figure shows binding across IKZF3, a known target gene of Helios (see our ChIP-qPCR figure).