Autophagy Essentials Antibody Sampler Kit
cat.: HAK21114
| 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. |
| Uniprot #: | SwissProt: Q14457 Human | O88597 Mouse | Q91XJ1 Rat | Q9GZQ8 Human | Q9CQV6 Mouse | Q62625 Rat | Q13501 Human | Q64337 Mouse | O08623 Rat | Q9H1Y0 Human | Q99J83 Mouse | Q3MQ06 Rat | P11279 Human | P11438 Mouse | P14562 Rat |
Images
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Fig1:
Western blot analysis of SQSTM1 / p62 on different lysates with Rabbit anti-SQSTM1 / p62 antibody (HA721171) at 1/10,000 dilution. Lane 1: HeLa cell lysate Lane 2: HeLa treated with 50μM Chloroquine for 18 hours cell lysate Lane 3: NIH/3T3 cell lysate Lane 4: C2C12 cell lysate Lane 5: PC-12 cell lysate Lysates/proteins at 20 µg/Lane. Predicted band size: 48 kDa Observed band size: 62 kDa Exposure time: 2 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 (HA721171) at 1/10,000 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. |
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Fig2:
Western blot analysis of SQSTM1 / p62 on different lysates with Rabbit anti-SQSTM1 / p62 antibody (HA721171) at 1/2,000 dilution. Lane 1: A549-si NT cell lysate Lane 2: A549-si SQSTM1 / p62 cell lysate Lysates/proteins at 10 µg/Lane. Predicted band size: 48 kDa Observed band size: 62 kDa Exposure time: 17 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 (HA721171) 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. |
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Fig3:
Immunocytochemistry analysis of C2C12 cells treated with or without 50μM Chloroquine for 24 hours labeling LC3B with Rabbit anti-LC3B antibody (ET1701-65) 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-LC3B antibody (ET1701-65) 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. |
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Fig4:
Western blot analysis of LC3B on different lysates with Rabbit anti-LC3B antibody (ET1701-65) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution. Lane 1: HeLa cell lysate Lane 2: HeLa treated with 50μM Chloroquine for 18 hours cell lysate Lane 3: C2C12 cell lysate Lane 4: C2C12 treated with 50μM Chloroquine for 18 hours cell lysate Lane 5: C6 cell lysate Lane 6: C6 treated with 50μM Chloroquine for 18 hours cell lysate Lane 7: mouse brain tissue lysate Lane 8: rat brain tissue lysate Lysates/proteins at 20 µg/Lane. Predicted band size: 14/16 kDa Observed band size: 14/16 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 (ET1701-65) at 1/2,000 dilution and competitor's antibody at 1/1,000 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. |
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Fig5:
Western blot analysis of LC3B on different lysates with Rabbit anti-LC3B antibody (ET1701-65) at 1/1,000 dilution. Lane 1: HCT 116 cell lysate Lane 2: HCT 116 treated with 50μM Chloroquine for 18 hours cell lysate Lane 3: U-87 MG cell lysate Lane 4: C6 cell lysate Lane 5: Mouse brain tissue lysate Lane 6: Rat brain tissue lysate Lysates/proteins at 20 µg/Lane. Predicted band size: 14/16 kDa Observed band size: 14/16 kDa Exposure time: 26 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 (ET1701-65) 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. |
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Fig6:
Immunocytochemistry analysis of C6 cells treated with or without 50μM Chloroquine for 24 hours labeling LC3B with Rabbit anti-LC3B antibody (ET1701-65) 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-LC3B antibody (ET1701-65) 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. |
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Fig7: Fluorescence multiplex immunohistochemical analysis of human gastric cancer (Formalin/PFA-fixed paraffin-embedded sections). Panel A: the merged image of anti-LC3B (ET1701-65, Green), anti-CD31 (M1511-8, Red) and anti-CA-IX (ET1701-51, Yellow) on human gastric cancer. HRP Conjugated UltraPolymer Goat Polyclonal Antibody HA1119/HA1120 was used as a secondary antibody. The immunostaining was performed with the Sequential Immuno-staining Kit (IRISKit™MH010101, www.luminiris.cn). The section was incubated in three rounds of staining: in the order of ET1701-65 (1/100 dilution), M1511-8 (1/2,000 dilution) and ET1701-51 (1/100 dilution) for 20 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 30 mins at 95℃. DAPI (blue) was used as a nuclear counter stain. Image acquisition was performed with Olympus VS200 Slide Scanner. |
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Fig8:
Western blot analysis of LAMP1 on different lysates with Rabbit anti-LAMP1 antibody (HA722827) at 1/1,000 dilution. Lane 1: Jurkat cell lysate Lane 2: HEK-293 cell lysate Lane 3: A431 cell lysate Lane 4: MCF7 cell lysate Lane 5: MDA-MB-231 cell lysate Lane 6: HeLa cell lysate Lane 7: HeLa cell lysate treated with deglycosylation Lane 8: NIH/3T3 cell lysate Lane 9: NIH/3T3 cell lysate treated with deglycosylation Lysates/proteins at 20 µg/Lane. Predicted band size: 44 kDa Observed band size: 120/44 kDa Exposure time: 21 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 (HA722827) 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. |
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Fig9:
Immunohistochemical analysis of paraffin-embedded mouse colon tissue with Rabbit anti-LAMP1 antibody (HA722827) at 1/10,000 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 (HA722827) at 1/10,000 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. |
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Fig10:
Immunofluorescence analysis of frozen mouse kidney tissue with Rabbit anti-LAMP1 antibody (HA722827) at 1/1,000 dilution. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for about 2 minutes in microwave oven. The tissues were blocked in 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (HA722827, green) at 1/1,000 dilution overnight at 4 ℃, washed with PBS. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclei were counterstained with DAPI (blue). |
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