FAM160B2 Antibody

What applications are validated for FAM160B2 antibodies in research?

FAM160B2 antibodies have been validated for multiple laboratory applications with specific dilution recommendations:

Different antibodies may perform optimally in different applications, so it's critical to select one validated for your specific experimental needs. For example, Proteintech's 17147-1-AP antibody has been specifically validated for detecting FAM160B2 in human brain tissue by Western blot and IHC .

What samples are recommended as positive controls for FAM160B2 antibody validation?

Based on the literature and product documentation, the following samples have been validated as positive controls:

When designing experiments, it's recommended to include these validated positive controls alongside experimental samples. For instance, mouse brain tissue has been confirmed to express detectable levels of FAM160B2 and serves as an excellent positive control for antibody validation .

How should researchers optimize Western blot protocols for FAM160B2 detection?

For optimal Western blot detection of FAM160B2:

• Sample preparation:

  • Use RIPA buffer with protease inhibitors for protein extraction

  • Load 20-40 μg of total protein per lane

  • Include positive controls such as human brain tissue or 293T cell lysate

• Gel electrophoresis and transfer:

  • Use 8-10% SDS-PAGE gels (FAM160B2 has a calculated MW of 82 kDa)

  • Transfer to PVDF membranes at 100V for 60-90 minutes

• Antibody incubation:

  • Block with 5% non-fat milk in TBST for 1 hour at room temperature

  • Dilute primary antibody 1:500-1:1000 for optimal signal-to-noise ratio

  • Incubate overnight at 4°C

• Detection optimization:

  • Use HRP-conjugated secondary antibodies at 1:5000-1:10000

  • Expected molecular weight: 82 kDa

  • Consider peptide competition assays to confirm specificity (as shown in ABIN6261667 data)

As noted in product validation data, a clear single band at approximately 82 kDa indicates specific detection, while multiple bands may suggest non-specific binding or protein degradation .

What is known about FAM160B2's function and role in cellular pathways?

FAM160B2 (also known as RAI16 or FHIP2B) has several emerging functional roles:

• Signaling pathway involvement:

  • Activates MAPK/ERK signaling pathways

  • Modulates TGFB signaling pathways

  • May function in retinoic acid-responsive pathways (suggested by the RAI16 synonym)

• Cellular localization and interactions:

  • Functions as part of the FHF complex (FHIP2B - FHF complex subunit HOOK interacting protein 2B)

  • Associates with microtubule-based transport systems

• Developmental roles:

  • May play a role in early lens development in mammals

  • Involved in cell growth and proliferation processes

• Tissue-specific functions:

  • Expression in brain and testis suggests specialized functions in these tissues

  • Upregulation in CD34+ cells in coronary artery disease patients indicates potential vascular roles

Research investigating these pathways typically requires combining antibody-based detection with functional assays such as pathway inhibition studies, protein-protein interaction analyses, or gene knockdown experiments .

How can researchers investigate FAM160B2's role in disease models?

Several approaches are recommended for studying FAM160B2 in disease contexts:

• Expression analysis in disease tissues:

  • Use IHC to examine FAM160B2 expression patterns in normal versus diseased tissues

  • Perform Western blot quantification across disease stages

• Patient-derived cell models:

  • Recent research used patient adipose stem cell (hASC)-derived adipocytes to study FAM160B2's role in diabetes

  • FAM160B2 was specifically induced by rosiglitazone (an antidiabetic drug) in certain patients

• Genetic variation studies:

  • Investigate how genetic variants affect FAM160B2 function

  • Evidence suggests genetic variation can influence drug responses through altered PPARγ binding near the FAM160B2 gene

• Research in neurological disorders:

  • FAM160B2 has been studied in relation to brain regions affected by Parkinson's disease, Lewy Body Dementia, and cognitive defects

  • Consider using FAM160B2 antibodies for comparative expression studies in these contexts

• Cancer research applications:

  • FAM160B2 (RAI16) enhances tumorigenesis in hepatocellular carcinoma

  • Study identified interactions between RAI16, PKA-RIIα, and 14-3-3θ in regulating HSP70 phosphorylation

For these advanced applications, combining antibody detection with genetic approaches and functional assays provides the most comprehensive analysis of FAM160B2's role in disease pathogenesis.

What techniques are recommended for studying protein-protein interactions involving FAM160B2?

To investigate FAM160B2's interactome and functional protein partnerships:

• Co-immunoprecipitation (Co-IP):

  • FAM160B2 antibodies have been validated for IP applications (0.5-4.0 μg antibody per 400-600 μg cell extracts)

  • Successfully used to identify interactions with PKA-RIIα and 14-3-3θ proteins

  • Protocol recommendations:

    • Use mild lysis buffers to preserve protein complexes

    • Pre-clear lysates to reduce non-specific binding

    • Include appropriate controls (IgG, input)

• Mass spectrometry following IP:

  • Identified interactions between RAI16 (FAM160B2) and heat shock protein 70

  • Recommended for unbiased discovery of novel interaction partners

• Proximity ligation assays:

  • Useful for confirming interactions in situ within cells

  • Requires antibodies from different host species against FAM160B2 and potential partners

• Functional validation:

  • Knockdown/knockout studies to confirm physiological relevance of interactions

  • Domain mapping experiments to identify critical regions for protein-protein binding

  • Mutational analysis to disrupt specific interactions

Recent research has used these approaches to demonstrate that RAI16 (FAM160B2) anchors both PKA-RIIα and 14-3-3θ, regulating heat shock protein 70 phosphorylation, with implications for cancer biology .

What are common issues in FAM160B2 detection and how can researchers overcome them?

Researchers may encounter several technical challenges when working with FAM160B2 antibodies:

• Multiple bands in Western blot:

  • Expected molecular weight is 82 kDa

  • Potential causes:

    • Protein degradation (add fresh protease inhibitors)

    • Post-translational modifications (consider phosphatase treatment)

    • Splice variants (verify with RT-PCR)

  • Validation: Use peptide competition assays as demonstrated with ABIN6261667

• Weak or no signal in IHC/IF:

  • Antigen retrieval is critical - some products specifically recommend TE buffer pH 9.0

  • Alternative approaches:

    • Try citrate buffer pH 6.0 as an alternative retrieval method

    • Optimize antibody concentration (1:50-1:200 range typically recommended)

    • Extend primary antibody incubation time (overnight at 4°C)

• Background or non-specific staining:

  • Increase blocking time and concentration (5% BSA or normal serum)

  • Optimize antibody dilution (start with manufacturer recommendations)

  • Consider using highly purified antibodies (affinity-purified products show better specificity)

• Cross-reactivity concerns:

  • Select antibodies validated across multiple applications

  • Include appropriate negative controls (non-expressing tissues/cells)

  • Consider knockout/knockdown validation when possible

When troubleshooting, always reference the specific validation data for your antibody, as performance can vary between products from different manufacturers.

How should researchers validate FAM160B2 antibody specificity in their experimental systems?

A comprehensive validation strategy includes:

• Positive and negative control samples:

  • Positive controls: human/mouse brain tissue, human testis, 293T cells

  • Negative controls: tissues with minimal FAM160B2 expression

  • Recombinant FAM160B2 protein can serve as an additional positive control

• Validation across multiple applications:

  • Confirm consistent results between WB, IHC, and IF when possible

  • Expected molecular weight (82 kDa) should be consistent in WB

• Blocking peptide competition:

  • Pre-incubate antibody with immunizing peptide

  • Should abolish specific signal as demonstrated in product validation images

• Genetic approaches:

  • siRNA/shRNA knockdown

  • CRISPR/Cas9 knockout

  • Overexpression systems

• Alternative antibodies:

  • Use multiple antibodies targeting different epitopes of FAM160B2

  • Compare results between different antibody clones

For example, antibody ABIN6261667 validation included Western blot analysis of recombinant protein with and without antigen-specific peptide competition to demonstrate specificity .

How is FAM160B2 research contributing to our understanding of disease mechanisms?

Recent studies have revealed several significant roles for FAM160B2 in disease processes:

• Metabolic disease research:

  • FAM160B2 was specifically induced by rosiglitazone in adipose stem cell-derived adipocytes from certain diabetes patients

  • PPARγ binding near the FAM160B2 gene was influenced by genetic variation, affecting drug response

  • This suggests FAM160B2 may be a key mediator in antidiabetic drug action and could contribute to personalized medicine approaches

• Cancer biology:

  • RAI16 (FAM160B2) enhances tumorigenesis in hepatocellular carcinoma

  • Functions as a novel tumor marker for hepatocellular carcinoma

  • Anchoring of both PKA-RIIα and 14-3-3θ by FAM160B2 regulates heat shock protein 70 phosphorylation, potentially affecting cancer cell survival

• Neurological disorders:

  • FAM160B2 expression has been studied in various brain regions affected by:

    • Parkinson's disease

    • Lewy Body Dementia

    • Cognitive defects

  • Different dysregulation patterns were observed in specific neuronal populations

• RNA methylation and synaptic function:

  • FAM160B2 has been studied in relation to N6-methyladenosine (m6A) RNA methylation

  • This research explores connections to synaptic function and local protein synthesis

  • May contribute to understanding pathophysiology of brain diseases

These diverse research areas demonstrate FAM160B2's importance across multiple disease contexts and suggest potential as a therapeutic target or biomarker.

What novel methodologies are being applied to study FAM160B2 function?

Cutting-edge approaches for FAM160B2 research include:

• Patient-derived cellular models:

  • Human adipose stem cell (hASC)-derived adipocytes used to study FAM160B2 in diabetes

  • Allows investigation of genetic variation effects on drug responses

• Integrated multi-omics approaches:

  • Combining RNA-seq, ChIP-seq, and proteomics to understand FAM160B2 regulation

  • Example: Study linking PPARγ binding sites to FAM160B2 expression in rosiglitazone response

• Advanced imaging techniques:

  • Subcellular localization studies using super-resolution microscopy

  • Co-localization with interaction partners using multi-color imaging

• Machine learning analysis:

  • Quantitative analysis of FAM160B2 expression in neurological disorders

  • Used to identify region-specific and cell type-specific dysregulation patterns

• CRISPR screening approaches:

  • FAM160B2 was identified in a genome-wide CRISPR screen related to cholesterol metabolism

  • Suggests potential roles in lysosomal trafficking and/or cholesterol homeostasis

These methodologies represent the frontier of FAM160B2 research and offer powerful approaches for understanding its functional roles in normal physiology and disease states.

What are emerging areas of FAM160B2 research that require further investigation?

Several promising research directions warrant further exploration:

• Precise molecular function clarification:

  • While FAM160B2 has been associated with multiple pathways (MAPK/ERK, TGFB), its exact molecular mechanisms remain incompletely understood

  • Structure-function studies would help define functional domains

• Role in metabolic diseases:

  • The specific induction of FAM160B2 by rosiglitazone in patient-derived adipocytes suggests important metabolic functions

  • Further research on its role in insulin sensitivity and glucose metabolism is needed

• Potential as a disease biomarker:

  • Preliminary evidence suggests potential as a tumor marker for hepatocellular carcinoma

  • Expression changes in neurological disorders warrant investigation as potential biomarkers

• Therapeutic targeting possibilities:

  • If critical for disease processes, FAM160B2 or its interaction partners could represent novel therapeutic targets

  • Drug discovery efforts focused on modulating FAM160B2 activity or expression

• Evolutionary conservation and comparative biology:

  • Studying FAM160B2 across species could provide insights into conserved functions

  • The protein is conserved across mammals, suggesting important biological roles

These research areas represent important knowledge gaps that, when addressed, will significantly advance our understanding of FAM160B2 biology and its relevance to human disease.

What are the recommended antibodies and protocols for specific experimental contexts?

Based on the available literature and validation data, the following recommendations can guide antibody selection:

When planning experiments, researchers should consider:

• The specific application requirements (WB, IHC, IF, IP)

• Species reactivity needs (human, mouse, rat)

• Validated positive controls available in their laboratory

• The specific epitope targeted by different antibodies

• Published literature using specific antibodies for similar applications