Flotillin-2 Recombinant Rabbit Monoclonal Antibody [PSH03-41]
cat.: HA721986
Product Type: Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity: Human, Mouse, Rat, Monkey
Applications: WB, IHC-P, IF-Cell, FC
Clonality: Monoclonal
Clone number: PSH03-41
Form: Liquid
Storage condition: Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer: PBS (pH7.4), 0.1% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration: 1ug/ul
Purification: Protein A affinity purified.
Molecular weight: Predicted band size: 47 kDa
Isotype: IgG
Immunogen: Recombinant protein within human Flotillin-2 aa 1-428 / 428.
Positive control: HeLa cell lysate, A431 cell lysate, COS-1 cell lysate, NIH/3T3 cell lysate, PC-12 cell lysate, mouse lung tissue lysate, rat lung tissue lysate, HeLa, 293T cell lysate, A549 cell lysate, MCF7 cell lysate, human lung tissue lysate, mouse skin tissue lysate, rat skin tissue lysate, NIH/3T3, PC-12, human breast cancer tissue, human liver cancer tissue, mouse lung tissue, rat lung tissue.
Subcellular location: Cell membrane, Membrane, caveola, Endosome, Membrane.
Recommended Dilutions:
  WB
  IHC-P
  IF-Cell
  FC
1:2,000-1:10,000
1:500-1:2,000
1:100
1:1,000
Uniprot #: SwissProt: Q14254 Human | Q60634 Mouse | Q9Z2S9 Rat
Alternative names: ECS-1 ECS1 Epidermal surface antigen 1 Epidermal surface antigen ESA ESA1 Flot2 FLOT2_HUMAN Flotillin 2 (epidermal surface antigen 1) Flotillin-2 M17S1 Membrane component chromosome 17 surface marker 1 Membrane component chromosome 17 surface marker 1 homolog membrane component, chromosome 17, surface marker 1 (35kD protein identified by monoclonal ECS-1) Membrane component, chromosome 17, surface marker 1 REG-1 Reggie-1 Reggie-2
Images
HA721986_1.jpg Fig1: Western blot analysis of Flotillin-2 on different lysates with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/5,000 dilution and competitor's antibody at 1/2,000 dilution.

Lane 1: HeLa cell lysate (15 µg/Lane)
Lane 2: A431 cell lysate (15 µg/Lane)
Lane 3: COS-1 cell lysate (15 µg/Lane)
Lane 4: NIH/3T3 cell lysate (15 µg/Lane)
Lane 5: PC-12 cell lysate (15 µg/Lane)
Lane 6: Mouse lung tissue lysate (30 µg/Lane)
Lane 7: Rat lung tissue lysate (30 µg/Lane)

Predicted band size: 47 kDa
Observed band size: 47 kDa

Exposure time: Lane 1-7 (left): 6 seconds; Lane 1-7 (right): 50 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 (HA721986) at 1/5,000 dilution and competitor's antibody at 1/2,000 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.
HA721986_2.jpg Fig2: Western blot analysis of Flotillin-2 on different lysates with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/10,000 dilution.

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

Lysates/proteins at 10 µg/Lane.

Predicted band size: 47 kDa
Observed band size: 47 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 (HA721986) at 1/10,000 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.
HA721986_3.jpg Fig3: Immunocytochemistry analysis of HeLa cells labeling Flotillin-2 with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/100 dilution.

Cells were fixed in 100% precooled methanol 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-Flotillin-2 antibody (HA721986) 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.
HA721986_4.jpg Fig4: Western blot analysis of Flotillin-2 on different lysates with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/2,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: 293T cell lysate
Lane 3: A549 cell lysate
Lane 4: A431 cell lysate
Lane 5: MCF7 cell lysate
Lane 6: COS-1 cell lysate
Lane 7: NIH/3T3 cell lysate
Lane 8: PC-12 cell lysate
Lane 9: Human lung tissue lysate
Lane 10: Mouse lung tissue lysate
Lane 11: Mouse skin tissue lysate (hot lysis)
Lane 12: Rat skin tissue lysate
Lane 13: Rat lung tissue lysate

Lysates/proteins at 30 µg/Lane.

Predicted band size: 47 kDa
Observed band size: 47 kDa

Exposure time: 20 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 (HA721986) 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.
HA721986_5.jpg Fig5: Immunocytochemistry analysis of NIH/3T3 cells labeling Flotillin-2 with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/100 dilution.

Cells were fixed in 100% precooled methanol 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-Flotillin-2 antibody (HA721986) 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.
HA721986_6.jpg Fig6: Immunocytochemistry analysis of PC-12 cells labeling Flotillin-2 with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/100 dilution.

Cells were fixed in 80% precooled methanol 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-Flotillin-2 antibody (HA721986) 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.
HA721986_7.jpg Fig7: Immunohistochemical analysis of paraffin-embedded human breast cancer tissue with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/2,000 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 (HA721986) 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.
HA721986_8.jpg Fig8: Immunohistochemical analysis of paraffin-embedded human liver cancer tissue with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/500 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 (HA721986) 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.
HA721986_9.jpg Fig9: Immunohistochemical analysis of paraffin-embedded mouse lung tissue with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/500 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 (HA721986) 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.
HA721986_10.jpg Fig10: Immunohistochemical analysis of paraffin-embedded rat lung tissue with Rabbit anti-Flotillin-2 antibody (HA721986) at 1/500 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 (HA721986) 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.
HA721986_11.jpg Fig11: Flow cytometric analysis of HeLa cells labeling Flotillin-2.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA721986, 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).
HA721986_12.jpg Fig12: Flow cytometric analysis of NIH/3T3 cells labeling Flotillin-2.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA721986, 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).
HA721986_13.jpg Fig13: Flow cytometric analysis of PC-12 cells labeling Flotillin-2.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA721986, 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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