TMEM131L Antibody, FITC conjugated

What is TMEM131L and what are its known functions?

TMEM131L (Transmembrane protein 131-like) is a membrane-associated protein that plays significant roles in cellular signaling pathways. Its primary functions include:

• Antagonizing canonical Wnt signaling by triggering lysosome-dependent degradation of Wnt-activated LRP6 (low-density lipoprotein receptor-related protein 6)

• Regulating thymocyte proliferation, particularly in immature single-positive thymocytes

• Acting as a negative regulator in the Wnt/β-catenin signaling pathway at the LRP6 coreceptor level

The protein is also known by alternative names including KIAA0922 .

What are the specifications of the TMEM131L Antibody, FITC conjugated?

The TMEM131L Antibody, FITC conjugated is characterized by the following specifications:

• Product type: Polyclonal antibody raised in rabbit

• Quantity: Typically supplied as 50μL

• Immunogen: Peptide sequence from human Transmembrane protein 131-like protein (amino acids 963-978)

• Reactivity: Human

• Isotype: IgG

• Conjugate: Fluorescein isothiocyanate (FITC)

• Purification method: Antigen affinity purified

• UniProt ID: A2VDJ0

• Gene ID: 23240 (as noted in some sources)

How should TMEM131L Antibody, FITC conjugated be stored and handled?

Proper storage and handling are critical for maintaining antibody functionality:

• Store at -20°C or -80°C upon receipt

• Avoid repeated freeze-thaw cycles as they can compromise antibody integrity and performance

• The antibody is supplied in liquid form containing 50% glycerol, 0.01M PBS (pH 7.4), and 0.03% Proclin 300 as a preservative

• For long-term storage, it is recommended to aliquot the antibody into smaller volumes before freezing to minimize freeze-thaw cycles

• Note that the product contains Proclin, which is considered hazardous and should be handled by trained staff only

What applications has the TMEM131L Antibody, FITC conjugated been validated for?

The TMEM131L Antibody, FITC conjugated has been validated for specific applications:

• Enzyme-Linked Immunosorbent Assay (ELISA)

• Immunofluorescence (IF) has been mentioned in some sources

• For other potential applications, it is recommended that researchers perform validation studies to determine optimal dilutions and conditions

How does TMEM131L regulate the Wnt signaling pathway, and how can this antibody help study this mechanism?

TMEM131L functions as an inhibitor of canonical Wnt signaling through a specific mechanism:

• It triggers lysosome-dependent degradation of the active phosphorylated form of LRP6 following Wnt activation

• The membrane-associated form of TMEM131L specifically targets the phosphorylated LRP6 for degradation while not affecting basal LRP6 expression

• Genetic mapping studies have revealed that this degradation depends on a conserved extracellular domain proximal to the membrane rather than the cytoplasmic part of TMEM131L

Researchers can use the TMEM131L Antibody, FITC conjugated to:

• Visualize the subcellular localization of TMEM131L using fluorescence microscopy

• Examine the co-localization of TMEM131L with LRP6 or other components of the Wnt signaling pathway

• Monitor changes in TMEM131L expression or distribution in response to Wnt pathway activation or inhibition

• Investigate the relationship between TMEM131L surface translocation and thymocyte development stages

What experimental considerations should be taken into account when using this antibody to study thymocyte proliferation?

When studying thymocyte proliferation using this antibody, researchers should consider:

• TMEM131L has been identified as a regulator of thymocyte proliferation, with its silencing resulting in hyperproliferation and developmental defects

• The antibody can be used to monitor stage-specific surface translocation of TMEM131L during thymopoiesis, which may be critical for regulating immature single-positive thymocyte proliferation arrest

• When designing experiments:

  • Include appropriate controls for FITC background fluorescence

  • Consider fixation methods that preserve both membrane integrity and fluorescence signal

  • Use complementary approaches to confirm findings, such as combining immunofluorescence with functional assays of Wnt signaling

  • When conducting flow cytometry experiments, use appropriate gating strategies to distinguish different thymocyte subpopulations

  • Consider time-course experiments to capture dynamic changes in TMEM131L expression during thymocyte development

How can contradictions in experimental results when using TMEM131L Antibody be resolved?

When facing contradictory results in experiments using TMEM131L Antibody, consider the following troubleshooting approaches:

• Verify antibody specificity:

  • Perform validation experiments using positive and negative controls

  • Consider using alternative antibodies targeting different epitopes of TMEM131L for confirmation

  • Include knockdown or knockout controls when possible

• Address technical variables:

  • Optimize antibody concentration through titration experiments

  • Test different fixation and permeabilization protocols, as membrane proteins can be sensitive to these procedures

  • Ensure proper blocking to reduce non-specific binding

  • Verify storage conditions have been maintained to preserve antibody activity

• Consider biological variables:

  • TMEM131L function appears to be context-dependent, with stage-specific roles during thymocyte development

  • The protein's inhibitory effect on Wnt signaling may vary depending on the activation state of the pathway

  • Surface translocation of TMEM131L may be a regulated event, affecting its detection in different cellular compartments

What are the considerations for designing experiments to study the interaction between TMEM131L and the LRP6 coreceptor?

When investigating TMEM131L-LRP6 interactions:

• Experimental approaches should account for the finding that TMEM131L specifically targets phosphorylated LRP6 for degradation following Wnt activation, rather than affecting baseline LRP6 expression

• Consider using the following methodologies:

  • Co-immunoprecipitation to detect physical interactions between TMEM131L and LRP6

  • Dual-labeling immunofluorescence with TMEM131L Antibody, FITC conjugated and antibodies against LRP6 (total and phosphorylated forms)

  • Live-cell imaging to track the dynamics of TMEM131L-mediated LRP6 degradation

  • Lysosomal inhibitors to confirm the degradation pathway

• Experimental design should include:

  • Time-course studies following Wnt pathway activation

  • Comparison of wild-type TMEM131L with mutants lacking the critical extracellular domain

  • Controls to distinguish between effects on total LRP6 versus phosphorylated LRP6

  • Quantification of both membrane-associated and total cellular TMEM131L

What methodological approaches can be used to investigate the role of TMEM131L in lysosomal degradation pathways?

To investigate TMEM131L's role in lysosomal degradation:

• Use TMEM131L Antibody, FITC conjugated in combination with lysosomal markers to assess co-localization using confocal microscopy

• Employ lysosomal inhibitors (e.g., bafilomycin A1, chloroquine) to determine if they prevent TMEM131L-mediated degradation of phosphorylated LRP6

• Design pulse-chase experiments to track the fate of phosphorylated LRP6 in the presence and absence of TMEM131L

• Consider subcellular fractionation followed by immunoblotting to quantify the distribution of TMEM131L and LRP6 in different cellular compartments

• For more comprehensive analysis, combine with proteomic approaches to identify additional TMEM131L-interacting proteins involved in lysosomal trafficking

What controls should be included when using TMEM131L Antibody, FITC conjugated in immunofluorescence studies?

For rigorous immunofluorescence experiments:

• Primary controls:

  • Isotype control: Use FITC-conjugated rabbit IgG at the same concentration to assess non-specific binding

  • Blocking peptide control: Pre-incubate the antibody with excess immunizing peptide (amino acids 963-978 of TMEM131L) to confirm specificity

  • Cells known to be negative for TMEM131L expression

• Secondary controls:

  • Autofluorescence control: Examine unstained samples to determine background fluorescence

  • Single-staining controls when performing multi-color immunofluorescence to set compensation parameters

  • Concentration gradient to determine optimal antibody dilution for maximum signal-to-noise ratio

How can TMEM131L Antibody be used to investigate the mechanism of Wnt signaling inhibition in different cell types?

To investigate TMEM131L's inhibitory effects on Wnt signaling across cell types:

• Compare TMEM131L expression and localization in cells with different responsiveness to Wnt signaling

• Combine antibody staining with TCF reporter assays to correlate TMEM131L expression with Wnt pathway activity

• Use the antibody to track TMEM131L expression following manipulation of the Wnt pathway (activation or inhibition)

• Design experiments that assess:

  • Cell-type specific differences in TMEM131L subcellular localization

  • Correlation between TMEM131L surface expression and β-catenin nuclear accumulation

  • The relationship between TMEM131L expression and phosphorylated LRP6 levels across cell types

This methodological approach can help determine whether TMEM131L's function as a Wnt inhibitor is universal or context-dependent.

How can TMEM131L Antibody be utilized in thymocyte development research?

For thymocyte development studies:

• Use flow cytometry with TMEM131L Antibody, FITC conjugated to:

  • Profile TMEM131L expression across different thymocyte subpopulations

  • Correlate TMEM131L levels with proliferation markers

  • Examine the relationship between TMEM131L expression and developmental stage markers

• Apply the antibody in ex vivo thymic organ culture systems to:

  • Track changes in TMEM131L expression during thymocyte maturation

  • Assess the impact of manipulating TMEM131L expression on developmental progression

  • Investigate the relationship between TMEM131L surface translocation and proliferation arrest

• Consider humanized mouse models where:

  • TMEM131L-deficient human CD34+ hematopoietic progenitors can be tracked for developmental abnormalities

  • The antibody can be used to confirm knockdown efficiency and study compensatory mechanisms

What methodological approaches can be used to study the structural requirements for TMEM131L function using this antibody?

To investigate structural requirements for TMEM131L function:

• Use the antibody to verify expression of TMEM131L mutants/truncations in structure-function studies

• Design experiments focusing on the conserved extracellular domain proximal to the membrane that has been implicated in phosphorylated LRP6 degradation

• Employ the following approaches:

  • Create domain deletion or point mutation constructs of TMEM131L

  • Verify mutant protein expression and localization using the antibody

  • Assess the ability of each mutant to inhibit Wnt signaling and promote LRP6 degradation

  • Correlate structural features with functional outcomes in thymocyte proliferation models