Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)

Rabbit Recombinant Monoclonal CBX1 / HP1 beta antibody. Suitable for IP, ChIP, WB, IHC-P, ICC/IF, Flow Cyt (Intra) and reacts with Mouse, Human, Rat samples.

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Images

Western blot - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (AB270988), expandable thumbnail

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

Reactivity data

Select an application

Product promiseTestedExpectedPredictedNot recommended

	IP	ChIP	WB	IHC-P	ICC/IF	Flow Cyt (Intra)
Human	Tested	Tested	Tested	Tested	Tested	Tested
Mouse	Tested	Expected	Tested	Tested	Tested	Tested
Rat	Expected	Expected	Tested	Tested	Expected	Expected

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/30	Notes -
Species Human	Dilution info 1/30	Notes -

Expected
Expected

Species	Dilution info	Notes
Species Rat	Dilution info Use at an assay dependent concentration.	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info 5 µg/mL	Notes -

Expected
Expected

Species	Dilution info	Notes
Species Mouse, Rat	Dilution info Use at an assay dependent concentration.	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/1000	Notes -
Species Rat	Dilution info 1/1000	Notes -
Species Human	Dilution info 1/1000	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/2000	Notes -
Species Rat	Dilution info 1/2000	Notes -
Species Human	Dilution info 1/2000	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/500	Notes -
Species Human	Dilution info 1/500	Notes -

Expected
Expected

Species	Dilution info	Notes
Species Rat	Dilution info Use at an assay dependent concentration.	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Mouse	Dilution info 1/5000	Notes -
Species Human	Dilution info 1/5000	Notes -

Expected
Expected

Species	Dilution info	Notes
Species Rat	Dilution info Use at an assay dependent concentration.	Notes -

Target data

See full target information CBX1

Function

Component of heterochromatin. Recognizes and binds histone H3 tails methylated at 'Lys-9', leading to epigenetic repression. Interaction with lamin B receptor (LBR) can contribute to the association of the heterochromatin with the inner nuclear membrane.

Alternative names

CBX, CBX1, Chromobox protein homolog 1, HP1Hsbeta, Heterochromatin protein 1 homolog beta, Heterochromatin protein p25, M31, Modifier 1 protein, p25beta, HP1 beta

Recommended products

Rabbit Recombinant Monoclonal CBX1 / HP1 beta antibody. Suitable for IP, ChIP, WB, IHC-P, ICC/IF, Flow Cyt (Intra) and reacts with Mouse, Human, Rat samples.

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Form: Liquid
Clonality: Monoclonal
Immunogen: The exact immunogen used to generate this antibody is proprietary information.
Clone number: EPR24242-78
Purification technique: Affinity purification Protein A
Concentration

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage duration: 1-2 weeks
Appropriate short-term storage conditions: +4°C
Appropriate long-term storage conditions: -20°C
Aliquoting information: Upon delivery aliquot
Storage information: Avoid freeze / thaw cycle

Notes

Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

This product is a recombinant monoclonal antibody, which offers several advantages including:

- High batch-to-batch consistency and reproducibility
- Improved sensitivity and specificity
- Long-term security of supply
- Animal-free batch production

For more information, read more on recombinant antibodies.

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.

Activity summary

CBX1 also known as HP1 beta is a chromatin-associated protein that plays an important role in chromatin organization and epigenetic regulation. This protein has a mass of approximately 22 kDa and localizes within the nucleus where it participates in heterochromatin formation. CBX1 recognizes and binds to histone H3 when it is methylated at lysine 9 (H3K9me3) a modification integral to forming tightly packed DNA regions. Expression of CBX1 occurs widely in many tissue types reflecting its fundamental role in genomic maintenance and cell cycle regulation.

Biological function summary

The CBX1 protein acts as an integral component of the chromatin remodeling complex. By binding to methylated histones CBX1 helps stabilize chromatin architecture and repress transcription of genes within heterochromatic regions. This function is essential for maintaining gene silencing which is critical for processes like development and differentiation. CBX1 collaborates with other HP1 isoforms such as HP1 alpha and HP1 gamma as part of the family of heterochromatin proteins that contribute collectively to chromatin dynamics and gene expression control.

Pathways

CBX1 plays a part in the epigenetic transcriptional repression and cell cycle regulation pathways. It associates with SUV39H1 a methyltransferase to recognize and bind the H3K9me3 mark adding stability to the methylation signal and contributing to gene silencing. Additionally CBX1 interacts with other proteins like DNMT1 a DNA methyltransferase reinforcing its role in sustaining repressive chromatin states and preserving genome integrity during DNA replication.

Associated diseases and disorders

CBX1 holds a significant association with cancer and neurological disorders. Aberrant expression or mutations in CBX1 can lead to disrupted chromatin states influencing oncogene activation or tumor suppressor gene silencing which are common events in cancer development. Furthermore altered CBX1 function links to neurological disorders where inappropriate chromatin remodeling contributes to disease pathology. Changes in the expression and interaction with other proteins including HP1 alpha highlight its potential role in disease mechanisms making CBX1 a relevant target for therapeutic research.

Product promise

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16 product images

Western blot - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Blocking and diluting buffer and concentration: Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS

False colour image of Western blot: Anti-CBX1/HP1 beta antibody [EPR24242-78] (ab270988) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution, shown in red.
In Western blot, ab270988 was shown to bind specifically to CBX1/HP1. A band was observed at 21 kDa in wild-type HAP1 cell lysates with no signal observed at this size in CBX1/HP1 beta knockout cell line cell. To generate this image, wild-type and CBX1/HP1 beta HAP1 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a PVDF-FL membrane. Membranes were blocked in Odyssey diluted in equal volume of 0.1 % TBS before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye^®680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/10000 dilution.
Performed under reducing conditions.
All lanes: Western blot - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988) at 1/1000 dilution
Lane 1: Wild-type HAP1 (human chronic myelogenous leukemia near-haploid cell) whole cell lysate at 20 µg
Lane 2: CBX1/HP1 beta knockout HAP1 whole cell lysate at 20 µg
Lane 3: 293T (human embryonic kidney epithelial cell), whole cell lysate at 20 µg
Lane 4: HeLa (human cervix adenocarcinoma epithelial cell), whole cell lysate at 20 µg
Lane 5: NIH/3T3 (mouse embryonic fibroblast), whole cell lysate at 20 µg
Secondary
Lanes 1 - 2: Goat Anti-Rabbit IgG H&L (IRDye®800CW) (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat Anti-Mouse IgG H&L (IRDye® 680RD) (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/10000 dilution
Lanes 3 - 5: Goat Anti-Rabbit IgG H&L (IRDye® 800CW) (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat Anti-Mouse IgG H&L (IRDye® 680RD) (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/10000 dilution
Predicted band size: 21 kDa
Western blot - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Blocking and diluting buffer and concentration: 5% NFDM/TBST
Exposure time: Lanes 1-4: 59 seconds Lanes 5-8: 3 minutes
Lysates/proteins at 10 μg per lane.
All lanes: Western blot - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988) at 1/1000 dilution
Lane 1: Mouse brain tissue lysate at 10 µg
Lane 2: Mouse heart tissue lysate at 10 µg
Lane 3: Mouse kidney tissue lysate at 10 µg
Lane 4: Mouse spleen tissue lysate at 10 µg
Lane 5: Rat brain tissue lysate at 10 µg
Lane 6: Rat heart tissue lysate at 10 µg
Lane 7: Rat spleen tissue lysate at 10 µg
Lane 8: Human kidney tissue lysate at 10 µg
Secondary
All lanes: Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/100000 dilution
Predicted band size: 21 kDa
Western blot - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Blocking and diluting buffer and concentration: 5% NFDM/TBST
Exposure time: Lanes 1-4: 15 seconds Lanes 5-7: 26 seconds
Lysates/proteins at 20 μg per lane.
All lanes: Western blot - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988) at 1/1000 dilution
Lane 1: MCF7 (human breast adenocarcinoma epithelial cell), whole cell lysate at 20 µg
Lane 2: A431 (human epidermoid carcinoma epithelial cell), whole cell lysate at 20 µg
Lane 3: ES-D3 (mouse embryonic mtipotent stem Cell), whole cell lysate at 20 µg
Lane 4: C2C12 (mouse myoblasts myoblast), whole cell lysate at 20 µg
Lane 5: C6 (rat glial tumor glial cell), whole cell lysate at 20 µg
Lane 6: RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage), whole cell lysate at 20 µg
Lane 7: PC-12 (rat adrenal gland pheochromocytoma), whole cell lysate at 20 µg
Secondary
All lanes: Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/100000 dilution
Predicted band size: 21 kDa
Immunocytochemistry/ Immunofluorescence - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized CBX1 KO HAP1 cells labelling CBX1/HP1 beta with ab270988 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody at 1/1000 dilution (Green).
Confocal image showing nuclear staining in parental HAP1 cell line, no staining in CBX1 KO HAP1 cell line is observed. Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Immunohistochemical analysis of paraffin-embedded Rat testis tissue labelling CBX1/HP1 beta with ab270988 at 1/2000 (0.281 ug/ml) followed by a ready to use Leica DS9800 (BOND^®, Polymer Refine Detection) was used. Nuclear staining on rat testis. The section was incubated with ab270988 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Leica DS9800 (BOND^®, Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins
Immunocytochemistry/ Immunofluorescence - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HeLa cells labelling CBX1/HP1 beta with ab270988 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing nuclear staining in HeLa cell line is observed. Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 dilution.
ChIP - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Chromatin was prepared from ES-D3 cells according to the Abcam Dual-X-ChIP protocol*. Cells were fixed with 1.5 mM EGS for 30mins and then formaldehyde for 10min.
The ChIP was performed with 25 μg of chromatin, 5 μg of ab270988 (red), or 5 μg of rabbit normal IgG Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730 (gray) and 25 μl of Protein A/G Dynabeads. The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

*http://www.abcam.com/resources?keywords=X%20ChIP%20protocol
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Immunohistochemical analysis of paraffin-embedded Mouse testis tissue labelling CBX1/HP1 beta with ab270988 at 1/2000 (0.281 ug/ml) followed by a ready to use Leica DS9800 (BOND^®, Polymer Refine Detection) was used. Nuclear staining on mouse testis. (PMID: 32651609) The section was incubated with ab270988 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Leica DS9800 (BOND^®, Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins
Immunoprecipitation - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
CBX1/HP1 beta was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 10ug with ab270988 at 1/30 dilution (2μg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab270988 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
Lane 1: HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate 10ug
Lane 2: ab270988 IP in HeLa whole cell lysate
Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab270988 in HeLa whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 15 seconds
All lanes: Immunoprecipitation - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Predicted band size: 21 kDa
Immunocytochemistry/ Immunofluorescence - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized NIH/3T3 cells labelling CBX1/HP1 beta with ab270988 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing nuclear staining in NIH/3T3 cell line is observed. Alexa Fluor® 594 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 dilution.
Immunoprecipitation - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
CBX1/HP1 beta was immunoprecipitated from 0.35 mg NIH/3T3 (mouse embryonic fibroblast), whole cell lysate 10 ug with ab270988 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab270988 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
Lane 1 (Input): NIH/3T3 (mouse embryonic fibroblast), whole cell lysate 10 ug
Lane 2: ab270988 IP in NIH/3T3 whole cell lysate
Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab270988 in NIH/3T3 whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 15 seconds
All lanes: Immunoprecipitation - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Predicted band size: 21 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Immunohistochemical analysis of paraffin-embedded Human skin tissue labelling CBX1/HP1 beta with ab270988 at 1/2000 (0.281 ug/ml) followed by a ready to use Leica DS9800 (BOND^® , Polymer Refine Detection) was used. Nuclear staining on human skin. (PMID: 25769025) The section was incubated with ab270988 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Leica DS9800 (BOND^® , Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Immunohistochemical analysis of paraffin-embedded Human testis tissue labelling CBX1/HP1 beta with ab270988 at 1/2000 (0.281 ug/ml) followed by a ready to use Leica DS9800 (BOND^®, Polymer Refine Detection) was used. Nuclear staining on human testis. The section was incubated with ab270988 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument Counterstained with Hematoxylin.
Secondary antibody only control: Secondary antibody is a ready to use Leica DS9800 (BOND^®, Polymer Refine Detection) was used.
Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 mins
Flow Cytometry (Intracellular) - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Flow cytometric analysis of 4% paraformaldehyde fixed, 90% methanol permeabilized HeLa (human cervix adenocarcinoma epithelial cell) cells. CBX1/HP1 beta with ab270988 at 1/5000 dilution (0.01ug)(Red) was compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor^® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.
Flow Cytometry (Intracellular) - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Flow cytometric analysis of of permeabilized NIH/3T3 (Mouse embryonic fibroblast cells (4% paraformaldehyde fixed, 90% methanol). Labelling CBX1/HP1 beta with ab270988 at 1/5000 dilution (0.01 ug)(Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor^® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.
Flow Cytometry (Intracellular) - Anti-CBX1 / HP1 beta antibody [EPR24242-78] (ab270988)
Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Wild-type HAP1 (human chronic myelogenous leukemia near-haploid cell, Right) / CBX1/HP1 beta knockout HAP1(Left) cells labelling CBX1/HP1 beta with ab270988 at 1/5000 dilution (0.01ug)(Red) compared with a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor^® 488, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081) at 1/2000 dilution was used as the secondary antibody.