Revision 1

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Store at -20C

IRF Family Antibody Sampler Kit

1 Kit

(9 x 20 microliters)

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Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Product Includes Product # Quantity Mol. Wt Isotype/Source
IRF-1 (D5E4) Rabbit Monoclonal Antibody847820 µl45-48 kDaRabbit IgG
IRF-2 Antibody494320 µl45 kDaRabbit
IRF-3 (D6I4C) Rabbit Monoclonal Antibody1190420 µl50-55 kDaRabbit IgG
IRF-4 (D9P5H) Rabbit Monoclonal Antibody1510620 µl51 kDaRabbit IgG
IRF-5 Antibody2026120 µl60 kDaRabbit
IRF-6 Antibody694820 µl58, 60 kDaRabbit
IRF-7 (D2A1J) Rabbit Monoclonal Antibody1301420 µl65 kDaRabbit IgG
IRF-8 (D20D8) Rabbit Monoclonal Antibody562820 µl50 kDaRabbit IgG
IRF-9 (D2T8M) Rabbit Monoclonal Antibody7668420 µl48 kDaRabbit IgG
Anti-rabbit IgG, HRP-linked Antibody7074100 µlGoat

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The IRF Family Antibody Sampler Kit contains reagents to examine the total protein levels of various IRF isoforms. The kit contains enough primary and secondary antibodies to perform two western blot experiments with each primary antibody.

Storage

Polyclonal antibodies are supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol, and monoclonal antibodies also contain less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibodies.

Background

Interferon regulatory factors (IRFs) comprise a family of transcription factors that function within the Jak/Stat pathway to regulate interferon (IFN) and IFN-inducible gene expression in response to viral infection (1). IRFs play an important role in pathogen defense, autoimmunity, lymphocyte development, cell growth, and susceptibility to transformation. The IRF family includes nine members: IRF-1, IRF-2, IRF-9/ISGF3γ, IRF-3, IRF-4 (Pip/LSIRF/ICSAT), IRF-5, IRF-6, IRF-7, and IRF-8/ICSBP. All IRF proteins share homology in their amino-terminal DNA-binding domains. IRF family members regulate transcription through interactions with proteins that share similar DNA-binding motifs, such as IFN-stimulated response elements (ISRE), IFN consensus sequences (ICS), and IFN regulatory elements (IRF-E) (2).

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Western blot analysis of extracts from control HeLa cells (lane 1) or IRF-3 knockout HeLa cells (lane 2) using IRF-3 (D6I4C) XP® Rabbit mAb, #11904 (upper) or β-actin (13E5) Rabbit mAb, #4970 (lower). The absence of signal in the IRF-3 knockout HeLa cells confirms specificity of the antibody for IRF-3.
undefined Image 1: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from HT-29 and G-361 cells, untreated (-) or treated with Human Interferon-α1 (hIFN-α1) #8927 (10 ng/ml, overnight; +), using IRF-7 (D2A1J) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
undefined Image 2: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using IRF-4 (D9P5H) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
undefined Image 3: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Western blot analysis of extracts from various cell lines using IRF-5 Antibody (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
undefined Image 4: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from SR, Ramos and A20 cells using IRF-2 Antibody.
undefined Image 5: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from Raji and Jurkat cells, untreated or treated with Human Interferon-α1 (hIFN-α1) #8927 (10 ng/ml; overnight) using IRF-8 (D20D8) Rabbit mAb.
undefined Image 6: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Western blot analysis of extracts from various cell lines using IRF-6 Antibody. Absence of IRF-6 in MCF7 cells has been reported previously (5). Doublet has been reported to result from phosphorylation (6).
undefined Image 7: IRF Family Antibody Sampler Kit
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
undefined Image 8: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from HeLa cells, untreated (-) or treated with Human Interferon-α1 (hIFN-α1) #8927 (10 ng/ml, 16 hr; +), using IRF-9 (D2T8M) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).
undefined Image 9: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Western blot analysis of HeLa cells, untreated or treated with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml, 4 hr), using IRF-1 (D5E4) XP® Rabbit mAb.
undefined Image 10: IRF Family Antibody Sampler Kit
CUT&RUN was performed with HeLa cells treated with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml, 4 hr) and IRF-1 (D5E4) 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 ERAP2, a known target gene of IRF-1 (see additional figure containing CUT&RUN-qPCR data).
undefined Image 11: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using IRF-3 (D6I4C) XP® Rabbit mAb.
undefined Image 12: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Western blot analysis of extracts from HT-29 cells transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence® IRF-7 siRNA I #13139 (+) or SignalSilence® IRF-7 siRNA II #13291 (+). Twenty-four hours after transfection, cells were treated with Human Interferon-α1 (hIFN-α1) #8927 (10 ng/ml, overnight; +) and analyzed by western blot using IRF-7 (D2A1J) Rabbit mAb #13014 (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower). The IRF-7 (D2A1J) Rabbit mAb confirms silencing of IRF-7 expression, while the β-Actin (D6A8) Rabbit mAb is used as a loading control.
undefined Image 13: IRF Family Antibody Sampler Kit
Immunoprecipitation of IRF-4 from RPMI 8226 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is IRF-4 (D9P5H) Rabbit mAb. Western blot analysis was performed using IRF-4 (D9P5H) Rabbit mAb.
undefined Image 14: IRF Family Antibody Sampler Kit
Immunoprecipitation of IRF-5 from HDLM-2 cell extracts. Lane 1 is 10% input, lane 2 is Normal Rabbit IgG #2729, and lane 3 is IRF-5 Antibody. Western blot analysis was performed using IRF-5 Antibody. Mouse Anti-Rabbit IgG (Light-Chain Specific) (D4W3E) mAb (HRP Conjugate) #93702 was used for detection.
undefined Image 15: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Western blot analysis of extracts from THP-1, RL, and A20 cell lines using IRF-8 (D20D8) Rabbit mAb.
undefined Image 16: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using IRF-9 (D2T8M) Rabbit mAb.
undefined Image 17: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using IRF-1 (D5E4) XP® Rabbit mAb.
undefined Image 18: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
CUT&RUN was performed with HeLa cells treated with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml, 4 hr) and IRF-1 (D5E4) 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 5 (upper), including ERAP2 (lower), a known target gene of IRF-1 (see additional figure containing CUT&RUN-qPCR data).
undefined Image 19: IRF Family Antibody Sampler Kit
Immunoprecipitation of IRF-3 from THP-1 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or IRF-3 (D6I4C) XP® Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using IRF-3 (D6I4C) XP® Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb #3678 and Anti-mouse IgG, HRP-linked Antibody #7076 were used as secondary antibodies.
undefined Image 20: IRF Family Antibody Sampler Kit
Western blot analysis of extracts from 293T cells, mock transfected or transfected with human and mouse IRF-8 constructs, using IRF-8 (D20D8) Rabbit mAb.
undefined Image 21: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

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Immunoprecipitation of IRF-9 from THP-1 cell extracts. Lane 1 is 10% input, lane 2 is with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is with IRF-9 (D2T8M) Rabbit mAb. Western blot analysis was performed using IRF-9 (D2T8M) Rabbit mAb.
undefined Image 22: IRF Family Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded mouse thymus using IRF-1 (D5E4) XP® Rabbit mAb performed on the Leica® BOND™ Rx.
undefined Image 23: IRF Family Antibody Sampler Kit
CUT&RUN was performed with HeLa cells treated with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml, 4 hr) and either IRF-1 (D5E4) 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 MOV10 intron 1 primers, human ERAP2 intron 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 to one.
undefined Image 24: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Confocal immunofluorescent analysis of HT-29 cells, untransfected (left) or transfected with Poly(I:C) (2.5 μg/ml, 6 hr; right), using IRF-3 (D6I4C) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
undefined Image 25: IRF Family Antibody Sampler Kit
Chromatin immunoprecipitations were performed with cross-linked chromatin from THP-1 and IRF-8 (D20D8) 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 GPR18/UBAC2, a known target gene of IRF8 (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product data sheet.
undefined Image 26: IRF Family Antibody Sampler Kit
Confocal immunofluorescent analysis of HeLa cells, untreated (left) or treated with Human Interferon-α1 (hIFN-α1) #8927 (10 ng/ml, 16 hr; right), using IRF-9 (D2T8M) Rabbit mAb (green). Actin filaments were labeled with DyLight™ 554 Phalloidin #13054 (red).
undefined Image 27: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

Cell Signaling Technology Logo
Imunohistochemical analysis of paraffin-embedded human breast carcinoma using IRF-1 (D5E4) XP® Rabbit mAb in the presence of control peptide (left) or antigen-specific peptide (right).
undefined Image 28: IRF Family Antibody Sampler Kit
Chromatin immunoprecipitations were performed with cross-linked chromatin from H929 cells and either IRF-4 (D9P5H) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human PRDM1 Exon 2 primers, SimpleChIP® Human SUB1 Promoter Primers #5156 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.
undefined Image 29: IRF Family Antibody Sampler Kit
Chromatin immunoprecipitations were performed with cross-linked chromatin from THP-1 and IRF-8 (D20D8) 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 13 (upper), including GPR18/UBAC2 (lower), a known target gene of IRF8 (see additional figure containing ChIP-qPCR data).
undefined Image 30: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

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Chromatin immunoprecipitations were performed with cross-linked chromatin from U266 cells treated with Human Interferon-α1 (hIFN-α1) #8927 (10 nM, 30 min) and either IRF-9 (D2T8M) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human MX1 Promoter Primers #57949, human WARS intron 1 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.
undefined Image 31: IRF Family Antibody Sampler Kit
Imunohistochemical analysis of paraffin-embedded human lymphoma using IRF-1 (D5E4) XP® Rabbit mAb.
undefined Image 32: IRF Family Antibody Sampler Kit
Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 THP-1 and either IRF-8 (D20D8) 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 human GPR18 promoter primers, SimpleChIP® Human MS4A7 Promoter Primers #9013, 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.
undefined Image 33: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

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Immunohistochemical analysis of paraffin-embedded mosue thymus using IRF-1 (D5E4) XP® Rabbit mAb.
undefined Image 34: IRF Family Antibody Sampler Kit
Imunohistochemical analysis of paraffin-embedded HeLa cell pellets, control (left) or IFN gamma-treated (right), using IRF-1 (D5E4) XP® Rabbit mAb.
undefined Image 35: IRF Family Antibody Sampler Kit
Confocal immunofluorescent analysis of HeLa cells, serum-starved (left) or treated with hIFNγ #8901 (100 ng/mL, 4 hr; right), using IRF-1 (D5E4) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin.
undefined Image 36: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

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Flow cytometric analysis of HeLa cells, untreated (blue) or treated with Human Interferon-γ (hIFN-γ) #8901 (green), using IRF-1 (D5E4) XP® Rabbit mAb.
undefined Image 37: IRF Family Antibody Sampler Kit
Simple Western™ analysis of lysates (1 mg/mL) from HeLa cells using IRF-3 (D6I4C) XP ® Rabbit mAb #11904. The virtual lane view (left) shows the target band (as indicated) at 1:50 and 1:250 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:50 (blue line) and 1:250 (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.
undefined Image 38: IRF Family Antibody Sampler Kit
Simple Western™ analysis of lysates (0.1 mg/mL) from HT-29 cells using IRF-7 (D2A1J) Rabbit mAb #13014. The virtual lane view (left) shows the target band (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.
undefined Image 39: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 1

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Simple WesternTM analysis of lysates (1.0 mg/mL) from THP-1 cells using IRF-8 (D20D8) Rabbit mAb #5628. The virtual lane view (left) shows the target band (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 JessTM Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
undefined Image 40: IRF Family Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.