Anti-HMGA2 antibody [RM1215]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HMGA2 antibody [RM1215] (AB320849)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HepG2 (human hepatocellular carcinoma epithelial cell) cells labelling HMGA2 with ab320849 at 1/5000 (0.10 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).

Confocal image showing nuclear staining in HepG2 cell line
(shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Low expression : HT-29 (PMID : 26964871).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Left Panel is applied with Anti-HMGA2 antibody [RM1215] (ab320849) at 1/5000 dilution (0.10 ug/ml);
Right panel is applied with Anti-HMGA2 antibody (ab97276) at 1/1000 dilution (0.08 ug/ml).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-HMGA2 antibody [RM1215] (AB320849)

HMGA2 was immunoprecipitated from 0.35 mg NCI-H1299 (human lung carcinoma epithelial cell) whole cell lysate with ab320849 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab320849 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : NCI-H1299 (human lung carcinoma epithelial cell) whole cell lysate 10ug
Lane 2 : ab320849 IP in NCI-H1299 (human lung carcinoma epithelial cell) whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab320849 in NCI-H1299 whole cell lysate

Blocking and dilution buffer and concentration : 5% NFDM/TBST

All lanes:

Immunoprecipitation - Anti-HMGA2 antibody [RM1215] (ab320849) at 1/30 dilution

All lanes:

NCI-H1299 (human lung carcinoma epithelial cell) whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

false

Exposure time: 145s

• WB

Supplier Data

Western blot - Anti-HMGA2 antibody [RM1215] (AB320849)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Low expression : HT-29 (PMID : 26964871)

The identity of the higher MW bands above 75 kDa are supposed to be non-specific binding.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-HMGA2 antibody [RM1215] (ab320849) at 1/1000 dilution

Lane 1:

NCI-H1299 (human lung carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 18 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-HMGA2 antibody [RM1215] (AB320849)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Lysate prepared in 1%SDS Hot lysis method was recommended.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

All lanes:

Western blot - Anti-HMGA2 antibody [RM1215] (ab320849) at 1/1000 dilution

Lane 1:

HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate , prepared in 1%SDS Hot lysis method at 20 µg

Lane 2:

HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate, prepared in RIPA lysis method at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 18 kDa,36 kDa

false

Exposure time: 180s