TENM1 Antibody, Biotin conjugated

What is TENM1 and why is it important in neuroscience research?

TENM1 (Teneurin-1) is a cell-surface adhesion protein belonging to the tenascin family and teneurin subfamily that plays critical roles in tissue development and axon guidance. The protein is also known as Ten-1, Protein Odd Oz/ten-m homolog 1, Tenascin-M1, and Teneurin transmembrane protein 1 .

TENM1 has an unusual structural architecture consisting of:

• A large cylindrical β-barrel with similarity to bacterial Tc-toxins

• A C-terminal toxin-like domain that emerges from the barrel

• An Ig-like domain that seals the bottom of the barrel

• A six-bladed β-propeller domain attached perpendicularly

This protein is primarily expressed in the developing central nervous system and may function as a cellular signal transducer. The TENM1 gene can undergo alternative splicing, resulting in multiple transcript variants that may have different functional properties . Research has shown that TENM1 forms high-affinity trans-cellular adhesion complexes with Latrophilins (LPHNs), a family of adhesion G protein-coupled receptors with roles in embryogenesis, tissue polarity, and synapse development .

What are the specifications of commercially available TENM1 Antibody, Biotin conjugated?

Commercial TENM1 Antibody, Biotin conjugated products typically have the following specifications:

How does biotin conjugation benefit detection of TENM1 in experimental settings?

Biotin conjugation provides several methodological advantages for detecting TENM1 in research applications:

Biotin (MW = 244.31 g/mol) is a small molecule that can be covalently attached to antibodies without significantly disturbing their natural function or binding specificity . The process of biotinylation enables:

• Enhanced sensitivity due to signal amplification through the biotin-streptavidin system, which has an extremely high affinity (Kd ≈ 10^-15 M)

• Versatile detection options using:

  • Anti-biotin antibodies

  • Streptavidin-tagged reporters (enzymes like HRP, fluorescent probes)

  • Avidin-conjugated detection systems

• Application flexibility in multiple techniques:

  • ELISA (both standard and capture formats)

  • Western blotting

  • Immunofluorescence microscopy

  • Immunohistochemistry

The biotin-streptavidin interaction is exploited in various research techniques to isolate and detect biotinylated molecules with high specificity and strong binding, making it particularly valuable for studying low-abundance neuronal proteins like TENM1 .

What are the optimal storage conditions for TENM1 Antibody, Biotin conjugated?

To maintain antibody activity and stability, the following storage conditions are recommended:

For reconstituted lyophilized antibodies, aliquoting is recommended before freezing to minimize freeze/thaw cycles. When using the antibody, allow it to equilibrate to room temperature and gently mix before use .

What makes TENM1's structural similarities to bacterial Tc-toxins significant for research?

The structural similarity between TENM1 and bacterial Tc-toxins provides important insights for research applications:

TENM1's extracellular region (ECR) shows striking structural similarity to bacterial Tc-toxins, with these key features:

• A large cylindrical β-barrel composed of a spiral of β-hairpins that partially encapsulates a C-terminal domain

• A dormant catalytic pocket similar to the active aspartyl protease domain in bacterial toxins, with notable differences:

  • TENM1 variant v1 has a D2345G alteration in one catalytic residue

  • Other TENMs maintain all catalytic residues, suggesting potential activity

  • Conservation of key residues (R2302 and D2315) that mediate catalytic activity in toxins

• Comparison of structural elements:

  • TENM1's six-bladed β-propeller resembles the TcA binding domain in bacterial component B

  • The Ig-like domain is structurally similar to receptor binding domains in Tc-toxin A-components

  • Unlike toxins, TENM1's β-propeller attaches perpendicularly to the barrel

This structural homology suggests evolutionary adaptation of a toxin-like structure for cell-cell communication functions, which has important implications for:

• Understanding TENM1 signaling mechanisms in neural development

• Potential proteolytic processing similar to bacterial toxins

• Designing experiments to target specific structural domains with appropriate antibodies

Researchers using TENM1 antibodies should consider these structural features when designing experiments, particularly when studying TENM1's interaction with binding partners or investigating potential proteolytic processing events.

How should researchers approach experimental design when studying TENM1-LPHN interactions?

When designing experiments to study TENM1-LPHN interactions using biotin-conjugated TENM1 antibodies, researchers should follow these methodological guidelines:

• Understand the interaction domains:

  • The TENM1/LPHN interaction is mediated by the lectin (LEC) and olfactomedin (OLF) domains of LPHN

  • The LEC domain contributes most of the binding affinity

  • The toxin-like region of TENM1 is critical for interaction with LPHN

• Experimental design considerations:

  Experimental Approach	Key Controls	Readout
  Flow cytometry	Empty vector controls Truncated TENM1 constructs (ΔTox and Δ4,5)	Biotinylated LEC binding
  Trans-cellular signaling	"Cis" setup (same cell population) "Trans" setup (different cell populations)	cAMP level measurement
  Binding affinity	Cis-dimerization mutation controls	Soluble LEC binding

• Critical methodological considerations:

  • Alternative splicing of TENM1 can act as a switch regulating trans-cellular adhesion to LPHN

  • One splice variant activates trans-cellular signaling (LPHN-dependent)

  • Another splice variant induces inhibitory postsynaptic differentiation

  • Biotin-conjugated antibodies must be selected to recognize the specific domain/splice variant of interest

• Signal transduction analysis:

  • LPHN mediates embryogenesis by modulating intracellular cAMP levels

  • TEN binding to LPHN induces trans-cellular cAMP signaling

  • Presence of TEN2 causes a decrease in cAMP levels in a LPHN-dependent manner

This approach allows researchers to properly evaluate TENM1-LPHN interactions while accounting for the complex structural and functional properties of these proteins.

What validation methods should be used to confirm TENM1 Antibody, Biotin conjugated specificity?

To ensure experimental validity, researchers should thoroughly validate the specificity of TENM1 Antibody, Biotin conjugated using these methodological approaches:

• Immunoassay validation:

  • Western blot: Verify band at expected molecular weight (280-305 kDa)

  • Be aware that the actual band may not be consistent with expectations due to post-translational modifications

  • Use verified positive control samples (e.g., U87-MG, U-251MG, HeLa for Western blot; C6, L929, U-2OS for immunofluorescence)

• Controls to include:

  • Positive control: Human brain tissue lysate (recommended)

  • Negative control: Samples lacking TENM1 expression

  • Blocking peptide competition: Pre-incubation with immunizing peptide should abolish signal

  • Secondary antibody only: To assess background signal

• Cross-reactivity assessment:

  • Test for cross-reactivity with other TENM family members (TENM2-4)

  • TENM1 antibody should not cross-react with other members of the TENM family

  • Verify species reactivity claims (Human, Mouse, Rat) with appropriate samples

• Biotin conjugation-specific validation:

  • Perform immunoelectrophoresis against anti-biotin, anti-host serum (e.g., anti-Rabbit), human IgM, and human serum

  • Should show single precipitin arc against expected targets

  • No reaction should be observed against unrelated proteins

• Functional validation:

  • For biotin-conjugated antibodies, confirm functional activity using streptavidin-based detection systems

  • Test in ELISA by assaying against target protein using appropriate dilution range (e.g., 1:3,000 to 1:17,000)

What are the potential issues when using biotin-conjugated antibodies in complex tissue samples?

Researchers working with TENM1 Antibody, Biotin conjugated in complex neural tissue samples should be aware of these potential challenges:

• Endogenous biotin interference:

  • Neural tissues contain endogenous biotin that can cause high background

  • Mitochondria-rich tissues may show higher biotin levels

  • Solution: Block endogenous biotin using streptavidin/avidin blocking steps prior to applying biotinylated antibodies

• Structure-activity relationship considerations:

  • Biotinylation requires modification of biotin's carboxylic acid to an amide or ester group

  • This modification affects biotin's interaction with uptake systems like sodium-dependent multivitamin transporter (SMVT)

  • Impact: May affect cellular uptake kinetics in live-cell experiments

• Lysine modification effects:

  • Biotin is attached to surface-exposed lysine residues on the antibody

  • Some antibodies may have lysine residues in their antigen-binding sites

  • Conjugation may affect binding activity and assay sensitivity

  • Solution: Validate each new lot of biotin-conjugated antibody against previous lots

• Detection system compatibility:

  • Different biotin conjugation chemistries are optimized for specific applications:

    • Type A: for assays using streptavidin-labeled detection reagents

    • Type B: for assays where biotinylated protein is captured by immobilized streptavidin

  • Select the appropriate biotin conjugation type for your experimental design

• Storage buffer interference:

  • Many biotin-conjugated antibodies contain glycerol and preservatives that may affect certain applications

  • Proclin 300 (0.03%) and high glycerol (50%) in storage buffers may interfere with some enzymatic applications

  • Solution: Consider buffer exchange for sensitive applications

How does alternative splicing of TENM1 impact antibody selection and experimental design?

Alternative splicing of TENM1 creates significant methodological challenges that researchers must address in experimental design:

• Splice variant implications:

  • TENM1 has several alternatively spliced transcript variants encoding different isoforms

  • Alternative splicing within the β-propeller domain acts as a functional switch

  • One splice variant activates LPHN-dependent trans-cellular signaling

  • Another variant induces inhibitory postsynaptic differentiation

• Antibody selection considerations:

  Experimental Goal	Antibody Selection Strategy	Methodological Consideration
  General TENM1 detection	Target conserved regions	Avoid regions affected by alternative splicing
  Specific splice variant detection	Target splice junctions	May require custom antibodies
  Functional domain studies	Target specific domains	Consider whether epitope is accessible in native conformation
  Protein interaction studies	Avoid binding/interaction domains	Antibody may interfere with protein-protein interactions

• Experimental design adaptations:

  • Include controls for different splice variants

  • Use multiple antibodies targeting different epitopes

  • Complement antibody studies with molecular techniques (RT-PCR, RNA-seq) to identify splice variants present in sample

  • When studying TENM1-LPHN interactions, consider how splice variants may affect results

• Interpretation considerations:

  • Different cell types/tissues may express different TENM1 splice variants

  • Developmental stage influences splice variant expression

  • Pathological conditions may alter splice variant ratios

  • Signal strength variations between samples may reflect splice variant differences rather than total TENM1 expression

What troubleshooting strategies should be employed for common issues with TENM1 Antibody, Biotin conjugated?

When encountering problems with TENM1 Antibody, Biotin conjugated experiments, researchers should implement these methodological troubleshooting approaches:

• High background in immunostaining:

  • Issue: Endogenous biotin or non-specific binding

  • Solution: Implement avidin/biotin blocking steps before primary antibody incubation

  • Alternative: Use lower antibody concentration (start with 1:200 for IF and titrate)

• Weak or no signal in Western blot:

  • Issue: Protein degradation or inefficient transfer of high molecular weight protein

  • Solution: Use lower percentage gels (6-8%) for better resolution of large proteins (280-305 kDa)

  • Alternative: Increase transfer time or use wet transfer for high molecular weight proteins

  • Note: Verify the observed MW may be around 280 kDa rather than the calculated 305 kDa due to protein modifications

• Multiple bands in Western blot:

  • Issue: Post-translational modifications or proteolytic processing

  • Solution: TENM1 can be cleaved into Ten-1 intracellular domain and Ten-1 extracellular domain

  • Note: TENM1 is cleaved into Ten-1 intracellular domain (IDten-1/Ten-1 ICD) and Teneurin C-terminal-associated peptide (TCAP-1/Ten-1 ECD)

• Storage-related activity loss:

  Issue	Likely Cause	Solution
  Gradual loss of activity	Repeated freeze/thaw cycles	Aliquot antibody upon receipt
  Sudden loss of activity	Improper shipping/handling	Store immediately at recommended temperature upon receipt
  Precipitate formation	Storage buffer incompatibility	Centrifuge product if not completely clear after standing at room temperature

• Inconsistent results between experiments:

  • Issue: Variation in biotin conjugation efficiency between lots

  • Solution: Use consistent lot when possible, or perform side-by-side validation of new lots

  • Alternative: Include standard/calibrator samples in each experiment to normalize results between runs

This troubleshooting guide addresses common technical challenges researchers face when working with biotin-conjugated TENM1 antibodies, providing practical solutions based on the molecular and biochemical properties of both TENM1 and biotin-conjugated detection systems.

How can researchers optimize detection systems for biotin-conjugated TENM1 antibodies?

Selecting the optimal detection system for biotin-conjugated TENM1 antibodies requires careful consideration of experimental goals and sample types:

• Enzymatic detection systems:

  Reporter System	Advantages	Best Applications
  Streptavidin-HRP	High sensitivity, stable signal	Western blot, IHC, ELISA
  Streptavidin-AP	Lower background in tissues with endogenous peroxidase	IHC of neural tissues
  Peroxidase Conjugated Streptavidin with ABTS	High sensitivity for ELISA	Quantitative ELISA (working dilution 1:3,000-1:17,000)

• Fluorescent detection optimization:

  • Streptavidin-fluorophore conjugates provide high sensitivity

  • Multiple fluorophore options enable multiplexing

  • For TENM1 in neural tissues, longer wavelength fluorophores reduce autofluorescence

  • Tyramide signal amplification can be used for low-abundance targets

• Signal amplification strategies:

  • Avidin-Biotin Complex (ABC) method enhances sensitivity

  • Tyramide Signal Amplification (TSA) provides 10-100× signal enhancement

  • Quantum dot-streptavidin conjugates offer high photostability and brightness

• Methodological considerations:

  • For weak signals: Implement biotin-free blocking reagents to reduce background

  • For multiplexing: Use sequential streptavidin-fluorophore incubations

  • For colocalization studies: Select fluorophores with minimal spectral overlap

  • For quantitative analysis: Include calibration standards in each experiment

• Application-specific optimizations:

  • For TENM1 in IF: Working dilution 1:50-1:200 provides optimal results

  • For TENM1 in WB: Working dilution 1:500-1:2000 is recommended

  • For ELISA: Pre-block plates with biotin-free blocking solution

These optimization strategies leverage the high-affinity biotin-streptavidin interaction while addressing the specific challenges of detecting TENM1 in complex biological samples.

What emerging research applications utilize TENM1 Antibody, Biotin conjugated?

Recent advances have expanded the applications of TENM1 Antibody, Biotin conjugated in neuroscience research:

• Neural circuit mapping:

  • TENM1's role in axon guidance makes it valuable for visualizing developing neural circuits

  • Biotin-conjugated antibodies enable multi-label experimental approaches

  • Co-labeling with synaptic markers reveals TENM1's role in synapse development

• Developmental neurobiology studies:

  • TENM1 is a direct target of the homeobox transcription factor EMX2

  • EMX2 is important for area specification in the developing cortex

  • Biotin-conjugated TENM1 antibodies can track expression changes during development

• Trans-synaptic signaling investigation:

  • TENM1-LPHN interactions mediate critical functions in CNS development

  • Biotin-conjugated antibodies enable visualization of trans-synaptic protein complexes

  • Can be combined with proximity ligation assays for in situ interaction detection

• Drug delivery targeting:

  • The biotin moiety can facilitate targeted drug delivery applications

  • Cancer cells often overexpress biotin uptake systems

  • Biotinylated antibodies can leverage this for enhanced cellular uptake (2-3 fold higher in some cancer cell lines)

• Structural biology applications:

  • The unique toxin-like architecture of TENM1 suggests potential proteolytic processing

  • Biotin-conjugated domain-specific antibodies can track potential cleavage products

  • May help elucidate whether TENM1 undergoes processing similar to bacterial toxins

These emerging applications leverage both the specificity of TENM1 antibodies and the versatility of biotin conjugation to address fundamental questions in neuroscience research.