PA5-32043

antibody from Invitrogen Antibodies

Targeting: LACTB2 CGI-83

Western blot (Supportive data in Antibodypedia)

Immunohistochemistry (Supportive data in Antibodypedia)

Other assay (Supportive data in Antibodypedia)

Antibody Data

Product number

PA5-32043

Provider

Invitrogen Antibodies

Product name

LACTB2 Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Recombinant protein fragment

Description

Recommended positive controls: A549, HCT116.

Concentration

0.61 mg/mL

References

Submitted references

Identification of LACTB2, a metallo-β-lactamase protein, as a human mitochondrial endoribonuclease.

Levy S, Allerston CK, Liveanu V, Habib MR, Gileadi O, Schuster G
Nucleic acids research 2016 Feb 29;44(4):1813-32

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western Blot using LACTB2 Polyclonal Antibody (Product # PA5-32043). Sample (30 µg of whole cell lysate). Lane A: A549. 12% SDS PAGE. LACTB2 Polyclonal Antibody (Product # PA5-32043) diluted at 1:1,000.

Immunohistochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunohistochemical analysis of paraffin-embedded human colon carcinoma, using LACTB2 Polyclonal Antibody (Product # PA5-32043) antibody at 1:500 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

Other assay

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Figure 1. LACTB2 is a mitochondrial, soluble and monomeric protein. ( A ) Mitochondria localization. HeLa cells were disrupted and the nuclear, cytoplasmic and mitochondria-containing fractions were separated and analyzed for the localization of LACTB2, using an immunoblotting assay. The nucleus-located protein, CPSF3L, was used as marker for nuclear proteins. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) served as a cytosolic marker and cytochrome C (Cyt C) as a mitochondrial marker. ( B ) Immunoblot analysis of mitochondrial extract following Proteinase K accessibility assay. TX-100, Triton X-100. PK, Proteinase K. PNPase is a mitochondrial protein located mainly in the intermembrane space and a small amount in the matrix. HSP60 is a mitochondrial matrix protein. Tom20 is a mitochondrial outer membrane protein. ( C ) LACTB2 is a soluble protein. Immunoblotting analysis of mitochondrial extract following alkaline sodium carbonate (Na 2 CO 3 ) extraction of soluble and peripheral membrane proteins. T, total extract. S, supernatant. P, pellet. Subunit 5 of the ATP synthase (ATP5) was used as a marker for intrinsic membrane proteins and Cyt C as a marker of soluble proteins. (D) Immunoblot analysis of the recombinant LACTB2 (Rec.) and the native protein in isolated mitochondria (Mit.) showing the same size on SDS-PAGE. Therefore, the mitochondria targeting signal is not cleaved upon entering the mitochondria and remain an integral part of the mature protein. ( E ) LACTB2 is

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Figure 8. Downregulation of LACTB2 expression resulted in modest accumulation of mitochondrial transcripts. ( A ) Quantitation of the amount of LACTB2 in the siRNA treated cells. siRNA duplex targeted for LACTB2 or scrambled siRNAs duplexes, as a negative control, were transfected into HEK-293 cells. Cells were collected 48 h post-transfection and the amount of LACTB2 was determined using immunoblotting assay. Proteins from equal number of cells were loaded in the lanes labeled 100%. Proteins from 50, 25 and 10% of the control and 150 and 200% of the siRNA LACTB2 treated cells were analyzed for the amount of LACTB2, in order to determine the degree of down-expression. ( B ) Medium acidification due to treatment with siRNA duplex, 48 h post-transfection. Ctrl--cells treated with scrambled siRNAs duplexes as a negative control. RNAi--cells treated with siRNA duplex targeted to LACTB2. ( C ) The amount of mitochondrial transcripts in the cells treated with the siRNA duplex targeted to LACTB2 (turquoise bars), or with scrambled siRNAs duplexes as a negative control (brown bars), was analyzed by qRT-PCR. The GAPDH transcript was used as the reference gene for normalization of the expression. The error bars display the standard errors of at least three independent experiments. ( D ) Northern blot analysis of HEK-293 cells that were not transfected (WT), transfected with scrambled siRNAs as control (Ctrl) or transfected with LACTB2 siRNA (RNAi). Hybridizations were performed with pr