HAK21109

Alzheimer's Disease Antibody Sampler Kit

cat.: HAK21109

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:	PBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Uniprot #:	SwissProt: P07196 Human \| P08551 Mouse \| P19527 Rat \| P10636 Human \| P10637 Mouse \| P19332 Rat \| P56817 Human \| P56818 Mouse \| P56819 Rat \| P49840 Human \| Q2NL51 Mouse \| P18265 Rat \| P49841 Human \| Q9WV60 Mouse \| P18266 Rat \| P10636-8 Human \| P10637 Mouse \| P19332 Rat \| P37840 Human \| O55042 Mouse \| P37377 Rat \| P05067 Human \| P12023 Mouse \| P05067 Human \| P12023 Mouse \| P08592 Rat

Images

Fig1: Western blot analysis of Amyloid Precursor Protein on different lysates with Rabbit anti-Amyloid Precursor Protein antibody (HA722139) at 1/1,000 dilution.

Lane 1: PC-3 cell lysate
Lane 2: K-562 cell lysate (negative)
Lane 3: Mouse brain tissue lysate
Lane 4: Mouse skeletal muscle tissue lysate (negative)
Lane 5: Rat brain tissue lysate
Lane 6: Rat skeletal muscle tissue lysate (negative)

Lysates/proteins at 20 µg/Lane.

Predicted band size: 87 kDa
Observed band size: 100 kDa

Exposure time: 20 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 (HA722139) 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: Western blot analysis of Phospho-Tau(S396) on SHSY5Y cell lysates.

Lane 1: SHSY5Y cells, whole cell lysate, 10ug/lane
Lane 2: SHSY5Y cells treated with 2.8ug/ul lambda-PP for 30 minutes, whole cell lysates, 10ug/lane

All lanes :
Anti-Phospho-Tau(S396) antibody (ET1611-68) at 1/500 dilution. Anti-GAPDH antibody (ET1601-4) at 1/10,000 dilution. Goat Anti-Rabbit IgG H&L (HRP) (HA1001) at 1/200,000 dilution.

Predicted band size: 79 kDa
Observed band size: 70/130 kDa

Blocking and diluting buffer: 5% BSA.

Exposure time: 2 minutes 34 seconds

Fig3: Western blot analysis of BACE1 on different lysates with Rabbit anti-BACE1 antibody (HA722244) at 1/1,000 dilution.

Lane 1: HeLa cell lysate (20 µg/Lane)
Lane 2: SH-SY5Y cell lysate (20 µg/Lane)
Lane 3: NIH/3T3 cell lysate (20 µg/Lane)
Lane 4: PC-12 cell lysate (20 µg/Lane)
Lane 5: Mouse brain tissue lysate (40 µg/Lane)
Lane 6: Rat brain tissue lysate (40 µg/Lane)

Predicted band size: 56 kDa
Observed band size: 56-80 kDa

Exposure time: 25 seconds; ECL: K1802;

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 (HA722244) 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.

Fig4: Immunofluorescence analysis of frozen mouse hippocampus tissue labeling Phospho-Tau (S396) with Rabbit anti-Phospho-Tau (S396) antibody (ET1611-68).
The tissues were blocked in 3% BSA for 30 minutes at room temperature, washed with PBS, and then probed with the primary antibody (ET1611-68, green) at 1/100 dilution overnight at 4℃, washed with PBS. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) was used as the secondary antibody at 1/200 dilution. Nuclei were counterstained with DAPI (blue). Image acquisition was performed with KFBIO KF-FL-400 Scanner.

Fig5: Western blot analysis of GSK3 alpha on different lysates with Rabbit anti-GSK3 alpha antibody (HA722217) at 1/2,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: A431 cell lysate
Lane 3: A549 cell lysate
Lane 4: Jurkat cell lysate
Lane 5: HCT 116cell lysate
Lane 6: HEK-293 cell lysate
Lane 7: U-87 MG cell lysate
Lane 8: MCF7 cell lysate
Lane 9: MDA-MB-231 cell lysate
Lane 10: SK-Br-3 cell lysate
Lane 11: Neuro-2a cell lysate
Lane 12: NIH/3T3 cell lysate
Lane 13: C6 cell lysate
Lane 14: COS-1 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 51 kDa
Observed band size: 51 kDa

Exposure time: 42 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 (HA722217) 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.

Fig6: Immunofluorescence analysis of frozen mouse hippocampus tissue labeling GSK3 beta with Rabbit anti-GSK3 beta antibody (ET1607-71).
The tissues were blocked in 3% BSA for 30 minutes at room temperature, washed with PBS, and then probed with the primary antibody (ET1607-71, green) at 1/100 dilution overnight at 4℃, washed with PBS. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) was used as the secondary antibody at 1/200 dilution. Nuclei were counterstained with DAPI (blue). Image acquisition was performed with KFBIO KF-FL-400 Scanner.

Fig7: Western blot analysis of GSK3 beta on different lysates with Rabbit anti-GSK3 beta antibody (ET1607-71) at 1/1,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: HEK-293 cell lysate
Lane 3: NIH/3T3 cell lysate
Lane 4: PC-12 cell lysate
Lane 5: Jurkat cell lysate
Lane 6: A549 cell lysate
Lane 7: HepG2 cell lysate
Lane 8: MCF7 cell lysate

Lysates/proteins at 15 µg/Lane.

Predicted band size: 47 kDa
Observed band size: 40 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 (ET1607-71) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.

Fig8: Western blot analysis of NF-L on different lysates with Rabbit anti-NF-L antibody (HA721538) at 1/1,000 dilution.

Lane 1: Mouse brain tissue lysate
Lane 2: Rat brain tissue lysate
Lane 3: Mouse hippocampus tissue lysate
Lane 4: Rat hippocampus tissue lysate
Lane 5: Human liver tissue lysate (negative)
Lane 6: Mouse liver tissue lysate (negative)
Lane 7: Rat liver tissue lysate (negative)

Lysates/proteins at 20 µg/Lane.

Predicted band size: 62 kDa
Observed band size: 68 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 (HA721538) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.

Fig9: Immunohistochemical analysis of paraffin-embedded human brain tissue with Rabbit anti-NF-L antibody (HA721538) at 1/1,000 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 (HA721538) at 1/1,000 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.

	Fig10: Immunohistochemical analysis of paraffin-embedded mouse hippocampus tissue with Rabbit anti-NF-L antibody (HA721538) at 1/1,000 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 (HA721538) at 1/1,000 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.
	Fig11: Immunocytochemistry analysis of mouse glial cells labeling NF-L with Rabbit anti-NF-L antibody (HA721538) at 1/1,000 dilution. Cells were fixed with 4% PFA (15 min), permeabilized with 0.25% TritonX-100 for 15 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4℃ with Rabbit anti-NF-L antibody (HA721538) at at 1/1,000 dilution. 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.
	Fig12: Immunofluorescence analysis of frozen mouse cerebellum tissue with Rabbit anti-NF-L antibody (HA721538) at 1/500 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 (HA721538, green) at 1/500 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).

Fig13: Western blot analysis of Tau on different lysates with Rabbit anti-Tau antibody (ET1612-44) at 1/1,000 dilution.

Lane 1: Mouse brain tissue lysate
Lane 2: Mouse hippocampus tissue lysate
Lane 3: Rat brain tissue lysate
Lane 4: Rat hippocampus tissue lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 79 kDa
Observed band size: 50-70 kDa

Exposure time: 1 minute 55 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 (ET1612-44) 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.

Fig14: Immunofluorescence analysis of frozen rat brain tissue with Rabbit anti-Tau antibody (ET1612-44) at 1/500 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 (ET1612-44, green) at 1/500 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).

Fig15: Immunofluorescence analysis of paraffin-embedded mouse cerebral cortex tissue labeling Tau with Rabbit anti-Tau antibody (ET1612-44) 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 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (ET1612-44, green) at 1/200 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).

Fig16: Immunofluorescence analysis of frozen rat cerebellum tissue with Rabbit anti-Alpha-Synuclein antibody (HA723035) at 1/500 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 (HA723035, green) at 1/500 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).

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