Anti-Methyltransferase like 5 antibody [EPR27864-16]

• IP

Supplier Data

Immunoprecipitation - Anti-Methyltransferase like 5 antibody [EPR27864-16] (AB319142)

Methyltransferase like 5 was immunoprecipitated from 0.35 mg HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate with ab319142 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab319142 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

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Lane 1 : HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : ab319142 IP in HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab319142 in HeLa whole cell lysate

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

All lanes:

Immunoprecipitation - Anti-Methyltransferase like 5 antibody [EPR27864-16] (ab319142) at 1/30 dilution

All lanes:

HeLa (human cervical adenocarcinoma 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

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Exposure time: 116s

• WB

Supplier Data

Western blot - Anti-Methyltransferase like 5 antibody [EPR27864-16] (AB319142)

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

Performed under reducing conditions.

In Western blot, ab319142 was shown to bind specifically to Methyltransferase like 5. Target of interest was observed at 25 kDa in wild-type HeLa cell lysates (lane 1) with no signal observed at this size in METTL5 knockout cell line (lane 2) (lane 2, knockout cell line ab265372 / knockout cell lysate ab258961).

The identity of the band at higher than 75 kDa (in lane 5) is unknown.

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

Exposure time : Lanes 1-3 : 59 seconds, lanes 4-5 : 180 seconds

All lanes:

Western blot - Anti-Methyltransferase like 5 antibody [EPR27864-16] (ab319142) at 1/1000 dilution

Lane 1:

Wild-type HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Lane 2:

METTL5 knockout HeLa whole cell lysate at 40 µg

Lane 3:

PC-3 (human prostate adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Lane 4:

HEK-293 (human embryonic kidney epithelial cell) whole cell lysate at 40 µg

Lane 5:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 40 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: 25 kDa,36 kDa

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