The TMEM88 antibody is a polyclonal or monoclonal antibody designed to detect the transmembrane protein 88 (TMEM88), a tumor suppressor implicated in various cancers. Key specifications include:
The TMEM88 antibody has been rigorously validated across multiple platforms:
The TMEM88 antibody has enabled critical insights into TMEM88’s role in cancer biology and therapeutic resistance:
Wnt/β-Catenin Signaling: TMEM88 inhibits Wnt signaling by interacting with Dishevelled-1 (Dvl-1), reducing β-catenin activity. Antibody-based studies confirm its role in suppressing oncogenic pathways in thyroid and ovarian cancers .
Subcellular Localization:
Cross-Reactivity: Validated for human, mouse, and rat; other species require confirmation .
Assay Optimization: Dilutions vary by application; e.g., 1 μg/mL for WB vs. 20 μg/mL for IF .
Clinical Translation: While TMEM88 is a prognostic biomarker, the antibody’s utility in routine diagnostics remains under investigation .
TMEM88 is a two-transmembrane-type protein that plays a critical role in regulating the Wnt signaling pathway. Its significance derives from its C-terminal tail that binds to the PDZ domain of Dishevelled (Dvl), a key component in Wnt signaling pathways . TMEM88 attenuates Wnt/β-catenin signaling induced by Wnt-1 ligand in a dose-dependent manner, while knockdown of TMEM88 by RNAi increases Wnt activity . This regulatory function makes TMEM88 an important research target in developmental biology, cancer research, and metabolic disease studies.
The human canonical protein consists of 159 amino acid residues with a molecular mass of 17.3 kDa . At least two isoforms of TMEM88 are known to exist, with most antibodies recognizing both isoforms .
TMEM88 exhibits complex subcellular localization patterns that are critical to understand when designing immunostaining experiments:
Plasma membrane: TMEM88 is prominently found at the cell surface, where it forms membrane-associated puncta with dynamic movements and internalization
Golgi apparatus: Significant expression in Golgi membranes has been consistently observed
Multivesicular bodies (MVBs): TMEM88 localizes to perinuclear MVBs that can be visualized as EEA1-positive structures resistant to digitonin permeabilization
Endocytic vesicles: Present in trafficking vesicles during internalization
This multi-compartment localization pattern suggests that TMEM88 trafficking between cellular compartments may be integral to its function. Importantly, nuclear versus cytoplasmic localization appears to correlate with different disease outcomes, particularly in breast cancer where cytoplasmic TMEM88 promotes tumor progression while nuclear TMEM88 may suppress tumors .
TMEM88 antibodies are utilized across multiple experimental applications:
While the calculated molecular weight of TMEM88 is 17.3 kDa, the observed molecular weight in Western blots is typically around 68 kDa , which may reflect post-translational modifications or protein complexes.
Sample preparation is critical for accurate detection of TMEM88 across its various subcellular localizations:
For MVB-specific detection:
Use digitonin permeabilization (before fixation) to remove cytosolic proteins while preserving membrane-associated TMEM88
Co-stain with the MVB marker EEA1, focusing on digitonin-resistant EEA1+ structures as a proxy for MVBs
Western blot analysis of digitonin-resistant versus soluble fractions can confirm MVB-associated TMEM88 (it co-segregates with membrane markers like NaK-ATPase)
For plasma membrane visualization:
Live imaging using total internal reflection fluorescence microscopy reveals membrane-associated puncta with dynamic movements
N-terminal GFP fusion constructs (GFP-TMEM88) are effective for tracking membrane localization and internalization dynamics
For complete subcellular profiling:
Immunoelectron microscopy using gold-conjugated anti-TMEM88 antibodies provides high-resolution localization data
Subcellular fractionation followed by Western blotting of distinct fractions helps quantify distribution patterns
These methodological approaches allow researchers to specifically track TMEM88 across different cellular compartments, which is essential for understanding its context-dependent functions.
When selecting TMEM88 antibodies, researchers should consider:
Epitope location: Antibodies generated against C-terminal epitopes (amino acids 110-160) are particularly effective for studying TMEM88 function, as this region contains the PDZ-binding domain critical for Wnt signaling regulation
Clonality considerations:
Polyclonal antibodies offer broader epitope recognition but may have batch-to-batch variation
Monoclonal antibodies provide consistent results but may miss certain conformational states
Isoform recognition: Verify whether the antibody recognizes all known TMEM88 isoforms if your research requires comprehensive detection
Application-specific validation:
Use of blocking peptides: When testing antibody specificity, include controls using the immunizing peptide to confirm binding specificity
The significant difference between calculated (17.3 kDa) and observed (approximately 68 kDa) molecular weights of TMEM88 in Western blots presents a methodological challenge . To address this discrepancy, researchers should consider:
Denaturation optimization:
Test different denaturation conditions (varying SDS concentrations, reducing agents, and heating temperatures)
Compare boiled versus non-boiled samples to detect potential heat-induced aggregation
Post-translational modification analysis:
Use phosphatase or glycosidase treatments prior to Western blotting to identify potential modifications
Apply mass spectrometry to characterize post-translational modifications that may affect migration
Cross-linking studies:
Use chemical cross-linking followed by Western blotting to determine if TMEM88 exists in stable complexes
Perform blue native PAGE to preserve protein complexes and compare with denaturing conditions
Alternative detection methods:
Validation with knockout/knockdown controls:
These methodological approaches can help distinguish between true TMEM88 signal and potential artifacts or complex formation.
TMEM88 regulates Wnt/β-catenin signaling through direct interaction with Dishevelled (Dvl) proteins via a specific C-terminal mechanism:
PDZ domain interaction:
The C-terminal tripeptide Val-Trp-Val (VWV) sequence of TMEM88 directly binds to the PDZ domain of Dvl
This interaction can be quantitatively measured using fluorescence spectroscopy, where Trp fluorescent polarization of the Val-Trp-Val tripeptide shows measurable changes upon Dvl-1 PDZ domain binding
Signaling inhibition mechanism:
TMEM88 binding promotes Wnt signalosome degradation in multivesicular bodies (MVBs)
In experimental systems, TMEM88 inhibits Siamois promoter-driven luciferase activity induced by Xdsh (Xenopus Dishevelled) but not by β-catenin, indicating it acts at the level of Dishevelled rather than downstream components
Context-dependent regulation:
Functional validation approaches:
These mechanistic insights explain how TMEM88 functions as a context-dependent regulator of Wnt signaling, rather than as a simple on/off switch.
TMEM88 exhibits striking context-dependent functions across different cancer types:
To properly study these context-dependent functions, researchers should employ:
Subcellular localization analysis:
Methylation analysis:
Pathway integration analysis:
Chemoresistance models:
Develop platinum-resistant cell lines to study TMEM88's role in drug resistance
Use patient-derived xenografts from treatment-resistant tumors
These methodological approaches help capture the complex and sometimes contradictory roles of TMEM88 across different cancer contexts.
TMEM88 exerts significant regulatory effects on lipid metabolism, particularly relevant in the context of Non-Alcoholic Fatty Liver Disease (NAFLD):
Regulation of key lipid metabolism factors:
Experimental approaches to study this function:
a) In vitro models:
Free Fatty Acid (FFA)-induced AML-12 hepatocyte cells transfected with pEGFP-C1-TMEM88 or TMEM88 siRNA
Quantification of lipid synthesis and metabolism markers using RT-qPCR and Western blotting
b) In vivo models:
Functional consequences:
TMEM88 accelerates the apoptotic rate of FFA-induced AML-12 cells (measurable via Annexin V-FITC/PI double staining flow cytometry)
TMEM88 inhibits proliferation of FFA-stimulated hepatocytes (detectable via EdU staining)
These effects may contribute to limiting the expansion of steatotic liver cells
Signaling pathway integration:
These findings suggest TMEM88 may represent a novel therapeutic target for metabolic disorders involving dysregulated lipid metabolism.
Investigating the dynamic trafficking of TMEM88 between cellular compartments requires specialized approaches:
Fluorescent protein fusion constructs:
Live cell imaging techniques:
Total Internal Reflection Fluorescence (TIRF) microscopy reveals membrane-associated puncta with dynamic movements and internalization events
Spinning disk confocal microscopy allows longer-term imaging with reduced phototoxicity
Dual-color imaging with markers for different compartments (e.g., Rab proteins) enables tracking of TMEM88 through the endosomal system
Photoactivatable/photoconvertible tags:
Fusion with photoconvertible fluorescent proteins (e.g., mEos, Dendra2) enables pulse-chase visualization of specific protein populations
This approach can determine the half-life of TMEM88 in different compartments and trafficking rates
Cargo trafficking assays:
Co-tracking with known endocytic cargo (transferrin, EGF) to determine if TMEM88 follows canonical trafficking routes
Pharmacological inhibitors of different trafficking pathways can identify the mechanisms of TMEM88 movement
FRAP (Fluorescence Recovery After Photobleaching) analysis:
Measure the mobility and exchange rates of TMEM88 in different compartments
Compare recovery kinetics between normal and disease conditions
These approaches provide complementary information about the complex trafficking patterns of TMEM88, which may be critical to its function in different cellular contexts.
The literature contains seemingly contradictory findings regarding TMEM88's function across different experimental systems. To address these contradictions, researchers should:
Systematically analyze context-dependent variables:
Develop standardized reporting:
Document complete experimental conditions including cell density, passage number, and media composition
Report antibody catalog numbers, dilutions, and validation methods
Specify exact genetic constructs used (full sequences preferred)
Combinatorial approaches:
Simultaneously manipulate TMEM88 and potential modifying factors
Use rescue experiments with wild-type vs. mutant constructs lacking specific domains
Employ dose-response studies rather than single-concentration experiments
Systems biology perspective:
Cross-validation between in vitro and in vivo systems:
Confirm cell line findings in primary cells and tissue samples
Develop tissue-specific conditional knockout models to address developmental compensation
These approaches can help resolve contradictory findings by identifying the specific conditions under which TMEM88 exerts different functions.
TMEM88's presence in multivesicular bodies (MVBs) suggests an important role in protein trafficking and degradation that requires specialized methodological approaches:
Advanced imaging techniques:
Super-resolution microscopy (STORM, PALM) to visualize TMEM88 distribution within MVB subdomains
Correlative light and electron microscopy (CLEM) to connect fluorescent signals with ultrastructural features
Live-cell imaging of GFP-TMEM88 with markers for MVB formation and maturation
Biochemical isolation of MVBs:
Density gradient fractionation to isolate MVB-enriched fractions
Immunomagnetic isolation using antibodies against MVB markers
Proteomic analysis of isolated MVBs to identify TMEM88-interacting proteins
Functional perturbation of MVB machinery:
ESCRT complex component knockdown/knockout to determine dependence of TMEM88 trafficking on canonical MVB formation
Manipulation of Rab proteins (particularly Rab7) to assess effects on TMEM88 localization
Lysosomal inhibition to distinguish between degradative and recycling MVB populations
Analysis of exosomal TMEM88:
Isolation of extracellular vesicles using ultracentrifugation or size-exclusion chromatography
Western blot and proteomic analysis of exosomal fractions for TMEM88 content
Functional studies of TMEM88-containing exosomes in recipient cells
MVB cargo sorting studies:
Investigation of TMEM88's potential role in sorting other proteins to MVBs
Analysis of ubiquitination patterns associated with TMEM88 trafficking
CRISPR-Cas9 screening for genes affecting TMEM88 MVB localization
These methodologies can help elucidate TMEM88's role in MVB biology, potentially revealing new functions in protein degradation, signaling regulation, and intercellular communication.
Based on current research, several approaches show promise for targeting TMEM88 therapeutically:
Context-specific targeting strategies:
Structural biology approaches:
Epigenetic modulation strategies:
Antibody-based therapeutics:
Pathway-based combination approaches:
Combining TMEM88 modulation with Wnt pathway inhibitors for synergistic effects
Rational design of combination therapies based on TMEM88's context-dependent functions
The development of these approaches requires overcoming current limitations, including insufficient structural information and limited understanding of TMEM88's full-length protein polypeptide and recognition epitopes .
Advanced genetic engineering techniques offer powerful new approaches to study TMEM88:
CRISPR-Cas9 genome editing applications:
Generation of endogenously tagged TMEM88 (with fluorescent proteins or affinity tags) to study physiological expression levels
Domain-specific mutations to dissect functional regions (e.g., transmembrane domains versus PDZ-binding motif)
Tissue-specific conditional knockout models to avoid developmental compensation
Single-cell analysis technologies:
Single-cell RNA-seq to identify cell populations with differential TMEM88 expression
Spatial transcriptomics to map TMEM88 expression patterns within complex tissues
Combined with lineage tracing to follow TMEM88 expression through developmental processes
Optogenetic and chemogenetic control:
Development of light-activated TMEM88 variants to achieve temporal control over its function
Chemically inducible dimerization systems to control TMEM88 interactions with binding partners
These approaches allow precise manipulation of TMEM88 function in specific subcellular compartments
Protein engineering approaches:
Creation of biosensors that report on TMEM88 conformational changes or binding events
Split-protein complementation assays to visualize TMEM88 interactions in living cells
Engineered TMEM88 variants with altered trafficking patterns to distinguish location-specific functions
Systems-level screening:
Genome-wide CRISPR screens to identify synthetic lethal interactions with TMEM88 in cancer contexts
Proteomic approaches like BioID or APEX proximity labeling to map the complete TMEM88 interactome
Metabolomic analysis following TMEM88 manipulation to understand broader metabolic consequences
These advanced approaches can provide unprecedented insights into TMEM88 biology that go beyond traditional methods, potentially revealing new functions and therapeutic opportunities.
Several methodological challenges must be addressed to effectively translate TMEM88 research findings:
Model system limitations:
Cell lines may not recapitulate the complex tissue environment and three-dimensional architecture
Patient-derived organoids or xenografts may better preserve tissue-specific TMEM88 functions
Development of physiologically relevant 3D culture systems incorporating stromal components
Technical standardization needs:
Standardized immunohistochemical protocols for consistent TMEM88 detection in clinical samples
Validated scoring systems for nuclear versus cytoplasmic TMEM88 expression
Development of companion diagnostics to identify patients likely to respond to TMEM88-targeted therapies
Mechanistic understanding gaps:
Translational research priorities:
Correlation of TMEM88 expression/localization patterns with clinical outcomes across larger patient cohorts
Development of biomarker panels combining TMEM88 with other prognostic markers
Identification of pharmacologically targetable nodes in TMEM88-related pathways
Drug development challenges:
Design of high-throughput screening assays for compounds that modulate TMEM88 function
Development of delivery systems that can target specific subcellular pools of TMEM88
Creation of therapeutic approaches that can distinguish between beneficial and harmful TMEM88 functions
Addressing these methodological challenges will be essential for effectively translating the growing body of TMEM88 research into clinical applications that improve patient outcomes.