HA751789

NFATc3 Recombinant Rabbit Monoclonal Antibody [PSH21-27]

cat.: HA751789

Product Type:	Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity:	Human, Mouse
Applications:	WB, IHC-P, IF-Cell, FC, IP
Clonality:	Monoclonal
Clone number:	PSH21-27

Form:	Liquid
Storage condition:	Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer:	1*PBS (pH7.4).
Concentration:	1ug/ul
Purification:	Protein A affinity purified.
Molecular weight:	Predicted band size: 116 kDa
Isotype:	IgG
Immunogen:	Recombinant protein within human NFATc3 aa 1-450.
Positive control:	Jurkat cell lysate, Daudi cell lysate, HeLa cell lysate, Raji cell lysate, NIH/3T3 cell lysate, RAW264.7 cell lysate, RAW264.7 treated with 10μg/mL LPS for 8 hours cell lysate, human thymus tissue, mouse thymus tissue, mouse stomach tissue, K-562, NIH/3T3.
Subcellular location:	Cytoplasm, Nucleus.
Recommended Dilutions: WB IHC-P IF-Cell FC IP	1:5,000 1:200 1:100 1:1,000 1-2μg/sample
Uniprot #:	SwissProt: Q12968 Human \| P97305 Mouse
Alternative names:	C80703 cytoplasmic 3 D8Ertd281e NF AT4 NF ATc3 NF-AT4 NF-ATc3 NFAC3_HUMAN NFAT4 NFATc3 NFATx Nuclear factor of activated T cells cytoplasmic 3 nuclear factor of activated T cells cytoplasmic calcineurin dependent 3 Nuclear factor of activated T-cells T cell transcription factor NFAT4 T-cell transcription factor NFAT4

Images

Fig1: Western blot analysis of NFATc3 on different lysates with Rabbit anti-NFATc3 antibody (HA751789) at 1/5,000 dilution.

Lane 1: Jurkat cell lysate
Lane 2: Daudi cell lysate
Lane 3: HeLa cell lysate
Lane 4: Raji cell lysate
Lane 5: NIH/3T3 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 116 kDa
Observed band size: 116 kDa

Exposure time: Lane 1-4: 20 seconds; Lane 5: 1 minute 33 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 (HA751789) 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.

Fig2: Western blot analysis of NFATc3 on different lysates with Rabbit anti-NFATc3 antibody (HA751789) at 1/5,000 dilution.

Lane 1: RAW264.7 cell lysate
Lane 2: RAW264.7 treated with 10μg/mL LPS for 8 hours cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 116 kDa
Observed band size: 116 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 (HA751789) 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.

	Fig3: Immunohistochemical analysis of paraffin-embedded human thymus tissue with Rabbit anti-NFATc3 antibody (HA751789) at 1/200 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 (HA751789) 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.
	Fig4: Immunohistochemical analysis of paraffin-embedded mouse thymus tissue with Rabbit anti-NFATc3 antibody (HA751789) at 1/200 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 (HA751789) 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.
	Fig5: Immunohistochemical analysis of paraffin-embedded mouse stomach tissue with Rabbit anti-NFATc3 antibody (HA751789) at 1/200 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 (HA751789) 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.

	Fig6: Immunocytochemistry analysis of K-562 cells labeling NFATc3 with Rabbit anti-NFATc3 antibody (HA751789) 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-NFATc3 antibody (HA751789) 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.
	Fig7: Flow cytometric analysis of K-562 cells labeling NFATc3. Cells were fixed and permeabilized. Then stained with the primary antibody (HA751789, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
	Fig8: Flow cytometric analysis of NIH/3T3 cells labeling NFATc3. Cells were fixed and permeabilized. Then stained with the primary antibody (HA751789, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

Fig9: NFATc3 was immunoprecipitated from 0.2 mg K-562 cell lysate with HA751789 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA751789 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: K-562 cell lysate (input)
Lane 2: HA751789 IP in K-562 cell lysate
Lane 3: Rabbit IgG instead of HA751789 in K-562 cell lysate

Blocking/Dilution buffer: primary antibody dilution (K1803)
Exposure time: 20 seconds; ECL: K1801

Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.