Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell lysate

5 Images

• WB

Lab

Western blot - Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell lysate (AB257270)

Lane 1 : Wild-type HeLa cell lysate (20 μg)

Lane 2 : FOLR1 knockout HeLa cell lysate (20 μg)

Lane 3 : JAR cell lysate (20 μg)

Lane 4 : MDA-MB-231 cell lysate (20 μg)

Lanes 1-4 : Merged signal (red and green). Green - ab221543 observed at 36, 38 kDa. Red - loading control, ab7291 observed at 52 kDa.

ab221543 Anti-Folate Binding Protein/FBP antibody [EPR20277] was shown to specifically react with Folate Binding Protein/FBP in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264921 (knockout cell lysate ab257270) was used. Wild-type and Folate Binding Protein/FBP knockout samples were subjected to SDS-PAGE. ab221543 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.

The molecular weight (36,38kDa) observed were knockout validated and consistent with what has been described in the literature (PMID : 23144806).

All lanes:

Western blot - Anti-Folate Binding Protein/FBP antibody [EPR20277] (<a href='/en-us/products/primary-antibodies/folate-binding-protein-fbp-antibody-epr20277-ab221543'>ab221543</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

FOLR1 knockout HeLa cell lysate at 20 µg

Lane 3:

JAR cell lysate at 20 µg

Lane 4:

MDA-MB-231 cell lysate at 20 µg

Predicted band size: 30 kDa

Observed band size: 38 kDa,36 kDa

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• WB

Unknown

Western blot - Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell lysate (AB257270)

Lane 1 : Wild-type HeLa cell lysate (20 μg)

Lane 2 : FOLR1 knockout HeLa cell lysate (20 μg)

Lane 3 : JAR cell lysate (20 μg)

Lane 4 : MDA-MB-231 cell lysate (20 μg)

Lanes 1-4 : Merged signal (red and green). Green - ab67422 observed at 38 kDa. Red - loading control, ab7291 observed at 52 kDa.

ab67422 Anti-Folate Binding Protein/FBP antibody was shown to specifically react with Folate Binding Protein/FBP in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264921 (knockout cell lysate ab257270) was used. Wild-type and Folate Binding Protein/FBP knockout samples were subjected to SDS-PAGE. ab67422 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 500 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Folate Binding Protein/FBP antibody (<a href='/en-us/products/primary-antibodies/folate-binding-protein-fbp-antibody-ab67422'>ab67422</a>) at 1/500 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

FOLR1 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-folr1-folate-binding-protein-fbp-knockout-hela-cell-line-ab264921'>ab264921</a>)

Lane 3:

JAR cell lysate at 20 µg

Lane 4:

MDA-MB-231 cell lysate at 20 µg

Predicted band size: 30 kDa

Observed band size: 38 kDa

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• Sanger seq

Unknown

Sanger Sequencing - Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell lysate (AB257270)

Allele-1 : 4 bp deletion in exon 1

• Sanger seq

Unknown

Sanger Sequencing - Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell lysate (AB257270)

Allele-2 : Insertion of the selection cassette in exon 1

• WB

Supplier Data

Western blot - Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell lysate (AB257270)

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

Negative control : K-562 (PMID : 15899836).

Lysates/proteins at 20 ug per lane.

In Western blot, ab322459 was shown to bind specifically to FOLR1. A band was observed at 37 kDa in wild-type HeLa cell lysates with whereas no signal observed at this size in FOLR1 knockout cell line ab264921(knockout cell lysate ab257270).

In Western blot, Anti-Vinculin antibody [EPR8185] (ab129002) staining at 1/10000 dilution.

All lanes:

Western blot - Anti-Folate Binding Protein/FBP antibody [RM1246] (<a href='/en-us/products/primary-antibodies/folate-binding-protein-fbp-antibody-rm1246-ab322459'>ab322459</a>) at 1/1000 dilution

Lane 1:

SK-OV-3 (human ovarian cancer epithelial cell) whole cell lysate at 20 µg

Lane 2:

JAR (human placenta choriocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

T-47D (human ductal breast epithelial tumor epithelial cell) whole cell lysate at 20 µg

Lane 4:

K-562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 5:

Wild-type HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 6:

Western blot - Human FOLR1 (Folate Binding Protein/FBP) knockout HeLa cell lysate (ab257270) at 20 µg

Lane 7:

Human ovary cancer tissue 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: 37 kDa,124 kDa

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Exposure time: 180s