NuRD Complex Antibody Sampler Kit
cat.: HAK21198
Product Type: Antibody Sampler Kit
Species reactivity: Human, Mouse
Applications: WB, IHC-P
Form: Liquid
Storage condition: Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer: 1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration: 1ug/ul
Uniprot #: SwissProt: Q12873 Human | Q14839 Human | Q6PDQ2 Mouse | Q13547 Human | O09106 Mouse | Q4QQW4 Rat | Q92769 Human | P70288 Mouse | F7ENH8 Rat | O95983 Human | Q9Z2D8 Mouse | F7EY92 Rat | Q13330 Human | Q8K4B0 Mouse | Q62599 Rat | Q16576 Human | Q60973 Mouse | Q71UF4 Rat
Entrez Gene: 216848 Mouse | 303241 Rat
Images
HAK21198_1.jpg Fig1: Western blot analysis of HDAC1 on different lysates with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/5,000 dilution and competitor's antibody at 1/1,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: HEK-293 cell lysate
Lane 3: MCF7 cell lysate
Lane 4: Jurkat cell lysate
Lane 5: L-929 cell lysate
Lane 6: C6 cell lysate

Lysates/proteins at 15 µg/Lane.

Predicted band size: 55 kDa
Observed band size: 65 kDa

Exposure time: 30 seconds; ECL: K1801;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1605-35) at 1/5,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% BSA at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21198_2.jpg Fig2: Immunocytochemistry analysis of HeLa cells labeling HDAC1 with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/500 dilution and competitor's antibody at 1/200 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/500 dilution and competitor's antibody at 1/200 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
HAK21198_3.jpg Fig3: Immunocytochemistry analysis of NIH/3T3 cells labeling HDAC1 with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/250 dilution and competitor's antibody at 1/200 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/250 dilution and competitor's antibody at 1/200 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
HAK21198_4.jpg Fig4: Immunocytochemistry analysis of C6 cells labeling HDAC1 with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
HAK21198_5.jpg Fig5: All lanes: Western blot analysis of HDAC1 with anti-HDAC1 antibody [SY12-04] (ET1605-35) at 1:1,000 dilution.
Lane 1: Wild-type HepG2 whole cell lysate (20 µg).
Lane 2: HDAC1 knockout HepG2 whole cell lysate (20 µg).

Predicted band size: 55 kDa
Observed band size: 65 kDa

ET1605-35 was shown to specifically react with HDAC1 in wild-type HepG2 cells. No band was observed when HDAC1 knockout samples were tested. Wild-type and HDAC1 knockout samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary Anti-HDAC1 antibody (ET1605-35, 1/1,000) and Anti-HSP90 antibody (ET1605-56, 1/10,000) were used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG H&L (HRP) Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.
HAK21198_6.jpg Fig6: Immunohistochemical analysis of paraffin-embedded mouse lymph nodes tissue with Rabbit anti-HDAC1 antibody (ET1605-35) at 1/800 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1605-35) at 1/800 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21198_7.jpg Fig7: HDAC1 was immunoprecipitated in 0.2mg HeLa cell lysate with ET1605-35 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1605-35 at 1/5,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: HeLa cell lysate (input)
Lane 2: ET1605-35 IP in HeLa cell lysate
Lane 3: Rabbit IgG instead of ET1605-35 in HeLa cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 43 seconds; ECL: K1802
HAK21198_8.jpg Fig8: Western blot analysis of HDAC2 on different lysates with Rabbit anti-HDAC2 antibody (ET1607-78) at 1/50,000 dilution and competitor's antibody at 1/1,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: HEK-293 cell lysate
Lane 3: MCF7 cell lysate
Lane 4: Jurkat cell lysate
Lane 5: L-929 cell lysate
Lane 6: NIH/3T3 cell lysate

Lysates/proteins at 15 µg/Lane.

Predicted band size: 55 kDa
Observed band size: 55 kDa

Exposure time: Lane 1-6 (left):30 seconds; Lane 1-6 (right): 1 minute 20 seconds; ECL: K1801;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1607-78) at 1/50,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21198_9.jpg Fig9: Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-HDAC2 antibody (ET1607-78) at 1/6,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1607-78) at 1/6,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21198_10.jpg Fig10: All lanes: Western blot analysis of HDAC2 with anti-HDAC2 antibody [SY29-02] (ET1607-78) at 1:1,000 dilution.
Lane 1: Wild-type HepG2 whole cell lysate (20 µg).
Lane 2: HDAC2 knockout HepG2 whole cell lysate (20 µg).

ET1607-78 was shown to specifically react with HDAC2 in wild-type HepG2 cells. No band was observed when HDAC2 knockout samples were tested. Wild-type and HDAC2 knockout samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary Anti-HDAC1 antibody (ET1607-78, 1/1,000) and Anti-HSP90 antibody (ET1605-56, 1/10,000) were used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG H&L (HRP) Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.
HAK21198_11.jpg Fig11: HDAC2 was immunoprecipitated from 0.2 mg NIH/3T3 cell lysate with ET1607-78 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1607-78 at 1/1,000 dilution. HRP Conjugated Anti-Rabbit IgG for IP Nano-secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: NIH/3T3 cell lysate (input)
Lane 2: ET1607-78 IP in NIH/3T3 cell lysate
Lane 3: Rabbit IgG instead of ET1607-78 in NIH/3T3 cell lysate

Blocking/Dilution buffer: primary antibody dilution (K1803)
Exposure time: 7 seconds; ECL: K1801
HAK21198_12.jpg Fig12: Western blot analysis of MTA1 on different lysates with Rabbit anti-MTA1 antibody (HA724208) at 1/5,000 dilution.

Lane 1: MCF7 cell lysate (20 µg/Lane)
Lane 2: Jurkat cell lysate (20 µg/Lane)
Lane 3: SW480 cell lysate (20 µg/Lane)
Lane 4: HeLa cell lysate (20 µg/Lane)
Lane 5: A549 cell lysate (20 µg/Lane)
Lane 6: 293T cell lysate (20 µg/Lane)
Lane 7: COS-1 cell lysate (20 µg/Lane)
Lane 8: Neuro-2a cell lysate (20 µg/Lane)
Lane 9: NIH/3T3 cell lysate (20 µg/Lane)
Lane 10: C6 cell lysate (20 µg/Lane)
Lane 11: Mouse bain tissue lysate (40 µg/Lane)

Predicted band size: 81 kDa
Observed band size: 80-85 kDa

Exposure time: 10 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA724208) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21198_13.jpg Fig13: Immunocytochemistry analysis of C6 cells labeling MTA1 with Rabbit anti-MTA1 antibody (HA724208) at 1/2,000 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-MTA1 antibody (HA724208) at 1/2,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
HAK21198_14.jpg Fig14: Immunohistochemical analysis of paraffin-embedded rat testis tissue with Rabbit anti-MTA1 antibody (HA724208) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA724208) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21198_15.jpg Fig15: MTA1 was immunoprecipitated from 0.2 mg Jurkat cell lysate with HA724208 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA724208 at 1/5,000 dilution. HRP Conjugated Anti-Rabbit IgG for IP Nano-secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: Jurkat cell lysate (input)
Lane 2: HA724208 IP in Jurkat cell lysate
Lane 3: Rabbit IgG instead of HA724208 in Jurkat cell lysate

Blocking/Dilution buffer: primary antibody dilution (K1803)
Exposure time: 3 seconds; ECL: K1801
HAK21198_16.jpg Fig16: Immunocytochemistry analysis of C6 cells labeling CHD3 with Rabbit anti-CHD3 antibody (HA720011) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-CHD3 antibody (HA720011) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
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