Human TMEM106B knockout A549 cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Lab
Western blot - Human TMEM106B knockout A549 cell line (AB273711)
Western blot : Anti-TMEM106B antibody staining at 0.1 µg/ml, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, the antibody was shown to bind specifically to TMEM106B. A band was observed at 45 kDa in wild-type A549 cell lysates with no signal observed at this size in TMEM106B knockout cell line ab273711 (knockout cell lysate ab273769). To generate this image, wild-type and TMEM106B knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
All lanes:
Anti-TMEM106B antibody staining at 0.1 µg/mL
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
Western blot - Human TMEM106B knockout A549 cell lysate (<a href='/en-us/products/cell-lysates/human-tmem106b-knockout-a549-cell-lysate-ab273769'>ab273769</a>) at 20 µg
Lane 3:
PANC-1 cell lysate at 20 µg
Lane 4:
Raji cell lysate at 20 µg
false
- NGS
Lab
Next Generation Sequencing - Human TMEM106B knockout A549 cell line (AB273711)
1 bp insertion after Arg88 of the WT protein
- NGS
Supplier Data
Next Generation Sequencing - Human TMEM106B knockout A549 cell line (AB273711)
Knockout achieved by CRISPR/Cas9; X = 1 bp insertion; Frameshift : 100%
- ICC/IF
Lab
Immunocytochemistry/ Immunofluorescence - Human TMEM106B knockout A549 cell line (AB273711)
Immunofluorescent analysis of 100% methanol-fixed, 0.1% TritonX-100 permeabilized Wildtype A549 (wildtype human lung carcinoma epithelial cell); TMEM106B KO A549 (TMEM106B knockout A549), ab273711 cells labelling TMEM106B with ab325555 at 1/500 (0.998 µg/ml) dilution , followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).
Confocal image showing lysosome staining in wildtype A549 cells (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
ab25631 Anti-LAMP2 mouse monoclonal antibody - Lysosome Marker was used to counterstain lysosomes at 1/200 dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 dilution (Magenta).
Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.
Complementary Products
Primary Antibodies
AB269860
Anti-IGJ antibody [EPR23130-113] - BSA and Azide free
BOND RX™ Validated|RabMAb|Recombinant
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IGJ antibody [EPR23130-113] - BSA and Azide free (AB269860)](https://content.abcam.com/products/images/igj-antibody-epr23130-113-bsa-and-azide-free-ab269860--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img82809.jpg)
- 0
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Reactivity data
Product details
We will provide viable cells that proliferate on revival.
This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
What's included?
Properties and storage information
Gene name
Gene editing type
Gene editing method
Knockout validation
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
Handling procedures
Initial handling guidelines
Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.
Subculture guidelines
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x104 cells/cm2 is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
- Do not allow the cell density to exceed 7x104 cells/cm2.
Culture medium
F-12K + 10% FBS
Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
TMEM106B is involved in several cellular processes impacting lysosome function and trafficking. It may become part of a protein complex that manages lysosomal positioning and pathway regulation within the cell. TMEM106B also affects cellular homeostasis and protein degradation which suggests a role in maintaining the proper balance of proteins and lipids within lysosomes. Alteration in TMEM106B expression or function can cause changes in lysosomal size and distribution indicating its role in cellular logistics.
Pathways
The TMEM106B protein becomes involved in the endolysosomal pathway and the autophagy-lysosome pathway. These pathways are critical for processing and breakdown of cellular waste and recycling of cellular components. In these pathways TMEM106B interacts with other proteins such as SORT1 (sortilin 1) and GRN (progranulin) both of which have important roles in protein sorting and trafficking. By influencing these pathways TMEM106B helps maintain cellular health and function.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male
Target data
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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