HA721925

HMGB2 Recombinant Rabbit Monoclonal Antibody [JE52-82]

cat.: HA721925

Product Type:	Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity:	Human, Mouse, Rat
Applications:	WB, IHC-P, IF-Cell, FC
Clonality:	Monoclonal
Clone number:	JE52-82

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
Purification:	Protein A affinity purified.
Molecular weight:	Predicted band size: 24 kDa
Isotype:	IgG
Immunogen:	Synthetic peptide within Human HMGB2 aa 1-50 / 209.
Positive control:	HeLa cell lysate, MCF7 cell lysate, HEK-293 cell lysate, K-562 cell lysate, NIH/3T3 cell lysate, PC-12 cell lysate, mouse testis tissue lysate, human breast carcinoma tissue, human thyroid tissue, mouse epididymis tissue, mouse testis tissue, rat testis tissue, MCF7, NIH/3T3, PC-12.
Subcellular location:	Chromosome. Cytoplasm. Nucleus.
Recommended Dilutions: WB IHC-P IF-Cell FC	1:1,000 1:500-1:2,000 1:100-1:250 1:1,000
Uniprot #:	SwissProt: P26583 Human \| P30681 Mouse \| P52925 Rat
Alternative names:	C80539 High mobility group (nonhistone chromosomal) protein 2 High mobility group box 2 High mobility group protein 2 High mobility group protein B2 HMG 2 HMG B2 HMG-2 HMG2 HMGB2 HMGB2_HUMAN

Images

Fig1: Western blot analysis of HMGB2 on different lysates with Rabbit anti-HMGB2 antibody (HA721925) at 1/1,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: MCF7 cell lysate
Lane 3: HEK-293 cell lysate
Lane 4: K-562 cell lysate
Lane 5: NIH/3T3 cell lysate
Lane 6: PC-12 cell lysate
Lane 7:Mouse testis tissue lysate

Cell lysates/proteins at 20 µg/Lane.
Tissue lysates/proteins at 30 µg/Lane.

Predicted band size: 24kDa
Observed band size: 28 kDa

Exposure time: 3 minutes 10 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 (HA721925) 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.

Fig2: Immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue with Rabbit anti-HMGB2 antibody (HA721925) at 1/500 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (HA721925) at 1/500 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.

	Fig3: Immunohistochemical analysis of paraffin-embedded human thyroid tissue with Rabbit anti-HMGB2 antibody (HA721925) at 1/500 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (HA721925) at 1/500 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 epididymis tissue with Rabbit anti-HMGB2 antibody (HA721925) at 1/2,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (HA721925) at 1/2,000 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.
	Fig5: Immunohistochemical analysis of paraffin-embedded mouse testis tissue with Rabbit anti-HMGB2 antibody (HA721925) at 1/2,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (HA721925) at 1/2,000 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.

	Fig6: Immunohistochemical analysis of paraffin-embedded rat testis tissue with Rabbit anti-HMGB2 antibody (HA721925) at 1/2,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (HA721925) at 1/2,000 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.
	Fig7: Immunocytochemistry analysis of MCF7 cells labeling HMGB2 with Rabbit anti-HMGB2 antibody (HA721925) 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-HMGB2 antibody (HA721925) 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.
	Fig8: Immunocytochemistry analysis of NIH/3T3 cells labeling HMGB2 with Rabbit anti-HMGB2 antibody (HA721925) 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-HMGB2 antibody (HA721925) 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.

Fig9: Immunocytochemistry analysis of PC-12 cells labeling HMGB2 with Rabbit anti-HMGB2 antibody (HA721925) at 1/250 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-HMGB2 antibody (HA721925) at 1/250 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.

Fig10: Flow cytometric analysis of MCF7 cells labeling HMGB2.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA721925, 1μg/mL) (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).

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