Niemann Pick C1 Recombinant Rabbit Monoclonal Antibody [JB87-33]
cat.: ET7107-57
Product Type: Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity: Human, Mouse, Rat
Applications: WB, IF-Cell, IF-Tissue, IHC-P, FC
Clonality: Monoclonal
Clone number: JB87-33
Form: Liquid
Storage condition: Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.
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: 142 kDa
Isotype: IgG
Immunogen: Recombinant protein within Human Niemann Pick C1 aa 1,160-1,278 / 1,278.
Positive control: HeLa cell lysate, HEK-293 cell lysate, Hep G2 cell lysate, NIH/3T3 cell lysate, C6 cell lysate, Mouse brain tissues lysate, Rat brain tissues lysate, human kidney tissue, rat kidney tissue, mouse testis tissue, SH-SY5Y.
Subcellular location: Endosome. Lysosome.
Recommended Dilutions:
  WB
  IF-Cell
  IF-Tissue
  IHC-P
  FC
1:500-1:2,000
1:50-1:200
1:50-1:200
1:50-1:200
1:50-1:100
Uniprot #: SwissProt: O15118 Human | O35604 Mouse
Entrez Gene: 266732 Rat
Alternative names: Niemann Pick C1 protein precursor Niemann Pick disease, type C1 Niemann-Pick C1 protein NPC NPC1 NPC1_HUMAN
Images
ET7107-57_1.jpg Fig1: Western blot analysis of Niemann Pick C1 on different lysates with Rabbit anti-Niemann Pick C1 antibody (ET7107-57) at 1/2,000 dilution.

Lane 1: HeLa (Human cervical adenocarcinoma cell) cell lysate (no heat) 20 µg/Lane
Lane 2: HEK-293 (Human embryonic kidney cell) cell lysate (no heat) 20 µg/Lane
Lane 3: Hep G2 (Human hepatocellular carcinoma cell) cell lysate (no heat) 20 µg/Lane
Lane 4: NIH/3T3 (Mouse fibroblast) cell lysate (no heat) 20 µg/Lane
Lane 5: C6 (Rat glioma cell) cell lysate (no heat) 20 µg/Lane
Lane 6: Mouse brain tissues lysate (no heat) 40 µg/Lane
Lane 7: Rat brain tissues lysate (no heat) 40 µg/Lane

Exposure time: 25 seconds; ECL: K1801

Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: ET7107-57, 1/2,000 in 5% NFDM/TBST, overnight at 4 ℃
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature

Predicted band size: 142.2 kDa
Observed band size: 190 kDa
ET7107-57_2.jpg Fig2: All lanes: Western blot analysis of Niemann Pick C1 with anti-Niemann Pick C1 antibody [JB87-33] (ET7107-57) at 1:1,000 dilution.
Lane 1: Wild-type HEK293 whole cell lysate (20 µg).
Lane 2: Niemann Pick C1 knockout HEK293 whole cell lysate (20 µg).

ET7107-57 was shown to specifically react with Niemann Pick C1 in wild-type HEK293 cells. No band was observed when Niemann Pick C1 knockout sample was tested. Wild-type and Niemann Pick C1 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 antibody (ET7107-57, 1/1,000) and Loading control antibody (Rabbit anti-HSP90, ET1605-56, 1/10,000) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG-HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.
ET7107-57_3.jpg Fig3: ICC staining of Niemann Pick C1 in A549 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET7107-57, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ET7107-57_4.jpg Fig4: ICC staining of Niemann Pick C1 in HepG2 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET7107-57, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ET7107-57_5.jpg Fig5: ICC staining of Niemann Pick C1 in PC-3M cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET7107-57, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ET7107-57_6.jpg Fig6: Immunohistochemical analysis of paraffin-embedded human kidney tissue using anti-Niemann Pick C1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-57, 1/50) for 30 minutes 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.
ET7107-57_7.jpg Fig7: Immunohistochemical analysis of paraffin-embedded rat kidney tissue using anti-Niemann Pick C1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-57, 1/50) for 30 minutes 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.
ET7107-57_8.jpg Fig8: Immunohistochemical analysis of paraffin-embedded mouse testis tissue using anti-Niemann Pick C1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-57, 1/50) for 30 minutes 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.
ET7107-57_9.jpg Fig9: Flow cytometric analysis of Niemann Pick C1 was done on SH-SY5Y cells. The cells were fixed, permeabilized and stained with the primary antibody (ET7107-57, 1/50) (red). After incubation of the primary antibody at room temperature for an hour, the cells were stained with a Alexa Fluor 488-conjugated Goat anti-Rabbit IgG Secondary antibody at 1/1,000 dilution for 30 minutes.Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.