Acetylation and deacetylation dynamics Antibody Sampler Kit
cat.: HAK21160
Product Type: Antibody Sampler Kit
Species reactivity: Human
Applications: WB
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: Q13547 Human | O09106 Mouse | Q4QQW4 Rat | O15379 Human | O88895 Mouse | Q6P6W3 Rat | Q96EB6 Human | Q923E4 Mouse | Q92830 Human | O95251 Human | Q5SVQ0 Mouse | Q810T5 Rat
Entrez Gene: 309757 Rat
Images
HAK21160_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.
HAK21160_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.
HAK21160_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.
HAK21160_4.jpg Fig4: 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.
HAK21160_5.jpg Fig5: 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
HAK21160_6.jpg Fig6: Western blot analysis of HDAC3 on different lysates with Rabbit anti-HDAC3 antibody (ET1610-5) at 1/2,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: K-562 cell lysate
Lane 3: HEK-293 cell lysate
Lane 4: HT-29 cell lysate
Lane 5: NIH/3T3 cell lysate
Lane 6: C6 cell lysate
Lane 7: COS-1 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 49 kDa
Observed band size: 49 kDa

Exposure time: 3 minutes; 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 (ET1610-5) at 1/2,000 dilution was 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.
HAK21160_7.jpg Fig7: Western blot analysis of HDAC3 on different lysates with Rabbit anti-HDAC3 antibody (ET1610-5) at 1/5,000 dilution.

Lane 1: HAP1-parental cell lysate
Lane 2: HAP1-HDAC3 KD cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 49 kDa
Observed band size: 49 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 (ET1610-5) at 1/5,000 dilution was used in 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.
HAK21160_8.jpg Fig8: Western blot analysis of KAT7 on different lysates with Rabbit anti-KAT7 antibody (ET1705-25) at 1/1,000 dilution.

Lane 1: HeLa cell lysate (20 µg/Lane)
Lane 2: NIH/3T3 cell lysate (20 µg/Lane)
Lane 3: RAW264.7 cell lysate (20 µg/Lane)
Lane 4: Mouse testis tissue lysate (40 µg/Lane)
Lane 5: Rat testis tissue lysate (40 µg/Lane)

Predicted band size: 71 kDa
Observed band size: 75 kDa

Exposure time: 3 minutes;

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 (ET1705-25) at 1/1,000 dilution was 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.
HAK21160_9.jpg Fig9: Immunohistochemical analysis of paraffin-embedded human tonsil tissue with Rabbit anti-HDAC3 antibody (ET1610-5) at 1/200 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 (ET1610-5) at 1/200 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.
HAK21160_10.jpg Fig10: Western blot analysis of SIRT1 on different lysates with Rabbit anti-SIRT1 antibody (ET1603-3) at 1/2,000 dilution.

Lane 1: HeLa cell lysate (15 µg/Lane)
Lane 2: HEK-293 cell lysate (15 µg/Lane)
Lane 3: A549 cell lysate (15 µg/Lane)
Lane 4: F9 cell lysate (15 µg/Lane)
Lane 5: Mouse testis tissue lysate (20 µg/Lane)
Lane 6: Rat testis tissue lysate (20 µg/Lane)

Predicted band size: 82 kDa
Observed band size: 110 kDa

Exposure time: 2 minutes 37 seconds;

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 (ET1603-3) at 1/2,000 dilution was 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.
HAK21160_11.jpg Fig11: Immunohistochemical analysis of paraffin-embedded rat testis tissue with Rabbit anti-SIRT1 antibody (ET1603-3) at 1/1,000 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 (ET1603-3) 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.
HAK21160_12.jpg Fig12: Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells with SIRT1 (ET1603-3) or Normal Rabbit IgG according to the ChIP protocol. The enriched DNA was quantified by real-time PCR using indicated primers. 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.
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