Human TREM2 Activity Antibody Sampler Kit
cat.: HAK21130
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: P43405 Human | P43405 Human | P43405 Human | P20138 Human | O43914 Human | Q9NZC2 Human
Images
HAK21130_1.jpg Fig1: Western blot analysis of Syk on different lysates with Rabbit anti-Syk antibody (HA721494) at 1/1,000 dilution.

Lane 1: Raji cell lysate
Lane 2: Daudi cell lysate
Lane 3: SW620 cell lysate
Lane 4: Ramos cell lysate

Lysates/proteins at 20 µg/Lane.
Predicted band size: 72 kDa
Observed band size: 70 kDa

Exposure time: 1 minute;
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 (HA721494) 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.
HAK21130_2.jpg Fig2: Western blot analysis of Phospho-Syk (Y525 + Y526) on different lysates with Rabbit anti-Phospho-Syk (Y525 + Y526) antibody (HA722034) at 1/1,000 dilution and competitor's antibody at 1/1,000 dilution.

Lane 1: Ramos cell lysate
Lane 2: Ramos serum starved for 16 hours add 12μg/mL human IgM (diluted in serum free medium) for 2 minutes cell lysate

Lysates/proteins at 20 µg/Lane.
Predicted band size: 72 kDa
Observed band size: 72 kDa
Exposure time: Lane 1-2 (left): 1 minute 2 seconds; Lane 1-2 (right): 2 minutes 18 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 (HA722034) at 1/1,000 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.
HAK21130_3.jpg Fig3: Western blot analysis of Phospho-Syk (Y352) on different lysates with Rabbit anti-Phospho-Syk (Y352) antibody (HA723362) at 1/5,000 dilution and pan Syk antibody at 1/1,000 dilution.

Lane 1: Ramos serum starved for 16 hours cell lysate
Lane 2: Ramos serum starved for 16 hours add 12μg/mL human IgM (diluted in serum free medium) for 2 minutes cell lysate
Lane 3: Ramos serum starved for 16 hours add 12μg/mL human IgM (diluted in serum free medium) for 2 minutes cell lysate, then the membrane treated with λpp for 1 hour

Lysates/proteins at 20 µg/Lane.
Predicted band size: 72 kDa
Observed band size: 72 kDa
Exposure time: 30 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 (HA723362) at 1/5,000 dilution and pan Syk 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.
HAK21130_4.jpg Fig4: Western blot analysis of CD33 on different lysates with Rabbit anti-CD33 antibody (HA721826) at 1/2,000 dilution.

Lane 1: THP-1 cell lysate
Lane 2: TF-1 cell lysate
Lane 3: HL-60 cell lysate
Lane 4: Jurkat cell lysate (negative)
Lane 5: SK-MEL-28 cell lysate (negative)

Lysates/proteins at 20 µg/Lane.
Predicted band size: 40 kDa
Observed band size: 70 kDa
Exposure time: 3 minutes; 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 (HA721826) 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.
HAK21130_5.jpg Fig5: Western blot analysis of CD33 on different lysates with Rabbit anti-CD33 antibody (HA721826) at 1/1,000 dilution.

Lane 1: THP-1 cell lysate
Lane 2: THP-1 cell lysate treated with deglycosylation

Predicted band size: 40 kDa
Observed band size: 40/70 kDa (Glycosylated)

Exposure time: 3 minutes; 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 (HA721826) 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.
HAK21130_6.jpg Fig6: Immunocytochemistry analysis of Ramos cells serum starved for 16 hours add 12μg/mL human IgM (diluted in serum free medium) for 2 minutes labeling Phospho-Syk (Y525 + Y526) with Rabbit anti-Phospho-Syk (Y525 + Y526) antibody (HA722034) at 1/1,000 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-Phospho-Syk (Y525 + Y526) antibody (HA722034) at 1/1,000 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.
HAK21130_7.jpg Fig7: Immunocytochemistry analysis of Ramos cells untreated / serum starved for 16 hours add 12μg/mL human IgM (diluted in serum free medium) for 2 minutes labeling Phospho-Syk (Y352) with Rabbit anti-Phospho-Syk (Y352) antibody (HA723362) at 1/1,000 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 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-Phospho-Syk (Y352) antibody (HA723362) at 1/1,000 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 (HA601187, 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.
HAK21130_8.jpg Fig8: Flow cytometric analysis of Ramos cells untreated (left) / serum starved for 16 hours add 12μg/mL human IgM (diluted in serum free medium) for 2 minutes (right) labeling Phospho-Syk (Y352).

Cells were fixed and permeabilized. Then stained with the primary antibody (HA723362, 1/1,000) (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).
HAK21130_9.jpg Fig9: Immunocytochemistry analysis of TF-1 (positive) and Jurkat (negative) labeling CD33 with Rabbit anti-CD33 antibody (HA721826) at 1/200 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-CD33 antibody (HA721826) 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.
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.