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STXBP1 KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout.

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Images

Key facts

Cell type

U-87 MG

Species or organism

Human

Tissue

Brain

Form

Liquid

Knockout validation

Sanger Sequencing, Western blot

Mutation description

Knockout.

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STXBP1 KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout.

Key facts

Cell type

U-87 MG

Form

Liquid

Mutation description

Knockout.

Disease

Glioblastoma

Concentration
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Properties

Gene name

STXBP1

Gene editing type

Knockout

Gene editing method

CRISPR technology

Knockout validation

Sanger Sequencing, Western blot

Zygosity

Homozygous

Cell culture

Biosafety level

EU: 1 US: 1

Viability

~ 80%

Adherent/suspension

Adherent

Gender

Male

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.

Culture medium

EMEM + 10% FBS

Cryopreservation medium

Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.

Storage

Shipped at conditions

Dry Ice

Appropriate short-term storage duration

A few days

Appropriate short-term storage conditions

-196°C

Appropriate long-term storage conditions

-196°C

Notes

Recommended control: Human wild-type U-87 MG cell line (ab278079). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.

Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.

Culture medium: EMEM + 10% FBS

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. Based on cell count, seed cells in an appropriate cell culture 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.
4. Incubate the culture at 37°C incubator with 5% CO2. 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.
  • A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.
  • Cells should be passaged when they have achieved 80-90% confluence.

We will provide viable cells that proliferate on revival.

This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.

Supplementary info

Biological function summary

This protein is important for the proper release of neurotransmitters into the synaptic cleft. Munc18-1 functions as a part of the neuronal exocytosis complex where it regulates the interaction with syntaxin-1 a core SNARE protein. This protein-protein interaction stabilizes an intermediate state necessary for vesicle priming and subsequent fusion ensuring efficient synaptic transmission. By controlling vesicle movement Munc18-1 influences communication between neurons.

Activity summary

Munc18-1 also known as Syntaxin-binding protein 1 is an important protein involved in the regulation of neurotransmitter release at synapses. It has a molecular mass of approximately 67 kDa. This protein is highly expressed in the brain where it plays a significant role in synaptic vesicle exocytosis. Munc18-1 interacts with components of the SNARE complex facilitating the docking and fusion of synaptic vesicles with the presynaptic membrane.

Pathways

Munc18-1 integrates into the neurotransmitter release pathway which is central to synaptic function. It closely associates with the SNARE complex involving proteins such as SNAP-25 and synaptobrevin. Another important pathway is the synaptic vesicle cycle where Munc18-1 plays a role in vesicle docking and priming stages. Through these interactions it ensures that synaptic vesicles are ready and able to release their contents upon stimulation.

Associated diseases and disorders

Alterations in Munc18-1 expression or function connect to neurological conditions like epilepsy and intellectual disability. In epilepsy dysregulation of neurotransmitter release due to modified Munc18-1 function disrupts synaptic communication leading to seizure activity. In the context of these disorders Munc18-1 interacts functionally with proteins such as VAMP2 contributing to the pathological state when disrupted. These interactions highlight the importance of Munc18-1 in maintaining normal neurological function.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

3 product images

  • Western blot - Human STXBP1 knockout U-87 MG cell line (ab322392), expandable thumbnail

    Western blot - Human STXBP1 knockout U-87 MG cell line (ab322392)

    ab124920 was shown to react with STXBP1 in wild-type U-87 MG cells in Western blot with loss of signal observed in STXBP1 knockout cell line ab322392. Wild-type U-87 MG and STXBP1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab124920 overnight at 4 °C at a 1/10000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

    These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

    All lanes: Western blot - Anti-Munc18-1 antibody [EPR4850] (AB124920) at 1/10000 dilution

    Lane 1: Wild-type U-87 MG lysate at 40 µg

    Lane 2: STXBP1 knock-out U-87 MG lysate at 40 µg

  • Immunocytochemistry/ Immunofluorescence - Human STXBP1 knockout U-87 MG cell line (ab322392), expandable thumbnail

    Immunocytochemistry/ Immunofluorescence - Human STXBP1 knockout U-87 MG cell line (ab322392)

    ab124920 was shown to react with STXBP1 in wild-type U-87 MG cells in immunocytochemistry with loss of signal observed in STXBP1 knockout cell line ab322392. Wild-type and knockout cells were mixed and pelleted at a 1:1 ratio on coverslips. The cells were fixed with 4% paraformaldehyde (15 min) then permeabilized with 0.1% Triton X-100 (10min) and then blocked with 1x PBS, 0.01% Triton X-100, 5% BSA, 5% NGS. The cells were then incubated with ab124920 at 1/100 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat anti-rabbit secondary antibody to (Alexa Fluor® 555) at 0.5 ?g/ml. Acquisition of the green (wild-type), red (antibody staining) and far-red (knockout) channels was performed. Representative grayscale images of the red channel are shown. Wild-type and knockout cells are outlined with yellow and magenta dashed line, respectively. Schematic representation of the mosaic strategy used is shown on the bottom-right panel. Image was acquired with a Zeiss(LSM-880).

    These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

  • Sanger Sequencing - Human STXBP1 knockout U-87 MG cell line (ab322392), expandable thumbnail

    Sanger Sequencing - Human STXBP1 knockout U-87 MG cell line (ab322392)

    55 bp deletion in Exon 3

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Product protocols

For this product, it's our understanding that no specific protocols are required. You can:

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

For licensing inquiries, please contact partnerships@abcam.com

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