AVPI1 Antibody

What is AVPI1 and what cellular functions does it participate in?

AVPI1 (arginine vasopressin induced 1) is a protein that may be involved in multiple cellular processes including MAP kinase activation, epithelial sodium channel (ENaC) down-regulation, and cell cycling . It has also been referred to as NPD013, PP5395, VIP32, and VIT32 in research literature . The human AVPI1 protein has a calculated molecular weight of approximately 21.5 kDa and is encoded by a gene located on chromosome 10 .

What experimental applications are AVPI1 antibodies validated for?

AVPI1 antibodies have been validated for several research applications:

For optimal results, each antibody should be titrated in your specific testing system to obtain optimal performance .

What species reactivity is available for AVPI1 antibodies?

Commercial AVPI1 antibodies have been developed with reactivity to multiple species:

• Human (most commonly available)

• Mouse

• Rat

• Cynomolgus/Rhesus macaque

• Gray mouse lemur (Microcebus murinus)

• Other mammals including feline, canine, bovine, and equine

When working with non-human samples, it is advisable to confirm cross-reactivity experimentally if not explicitly validated by the manufacturer.

How can I validate the specificity of an AVPI1 antibody in my experimental system?

A comprehensive validation approach should include multiple strategies:

• Positive controls: Use tissues known to express AVPI1, such as human lung cancer tissue which has been documented to show positive IHC staining

• Overexpression validation: Compare staining in cells transfected with AVPI1 expression vectors versus control-transfected cells. HEK293T cells have been successfully used for this purpose

• Western blot analysis: Confirm that the antibody detects a band of the expected molecular weight (approximately 21.5 kDa for human AVPI1)

• Peptide competition: Pre-incubate the antibody with the immunizing peptide to block specific binding

• Knockout/knockdown validation: Compare staining in wild-type cells versus AVPI1 knockout or knockdown cells

What are the optimal conditions for AVPI1 immunohistochemistry?

Based on published protocols, the following conditions have been recommended:

• Antigen retrieval: Use TE buffer at pH 9.0 as the preferred method. Alternatively, citrate buffer at pH 6.0 may also be effective

• Antibody dilution: Start with a dilution range of 1:50-1:500 and optimize for your specific tissue type

• Detection system: Both chromogenic (HRP-based) and fluorescent detection systems have been successfully used

• Controls: Include positive tissue controls (e.g., human lung cancer tissue) and appropriate negative controls

How can I design experiments to investigate the relationship between AVPI1 and MAP kinase signaling?

Given that AVPI1 may be involved in MAP kinase activation , consider these experimental approaches:

• Co-immunoprecipitation studies: Identify physical interactions between AVPI1 and components of MAP kinase cascades

• Phosphorylation analysis: After AVPI1 knockdown or overexpression, assess phosphorylation status of MAP kinase pathway proteins (ERK1/2, p38, JNK) using phospho-specific antibodies

• Domain mapping: Generate truncated or mutated versions of AVPI1 to identify regions critical for MAP kinase regulation

• Stimulation experiments: Expose cells to growth factors or stressors known to activate MAP kinase pathways, then analyze changes in AVPI1 expression, localization, or post-translational modifications

• Proximity ligation assay: Detect in situ protein-protein interactions between AVPI1 and MAP kinase components

What is known about AVPI1 expression in cancer tissues?

AVPI1 has been detected in human cancer tissues, with documented expression in lung cancer specimens using immunohistochemistry . The Human Protein Atlas provides expression data across multiple cancer types . When studying AVPI1 in cancer contexts, consider:

• Comparing expression levels between tumor and matched normal adjacent tissue

• Correlating expression with clinicopathological parameters

• Investigating potential prognostic significance

• Examining co-expression with related signaling molecules

What antigen retrieval methods optimize AVPI1 detection in fixed tissues?

The choice of antigen retrieval method can significantly impact AVPI1 detection in immunohistochemistry:

• Heat-induced epitope retrieval (HIER):

  • Primary recommendation: TE buffer at pH 9.0

  • Alternative method: Citrate buffer at pH 6.0

• Protocol optimization:

  • Heating time: Typically 10-20 minutes

  • Temperature: 95-100°C

  • Cooling: Slow return to room temperature

  • Buffer volume: Ensure tissue sections are completely immersed

Compare both methods using control tissues with known AVPI1 expression to determine which provides the highest signal specificity and lowest background for your specific tissue type.

How can I troubleshoot weak or inconsistent AVPI1 signal in Western blot?

When experiencing weak or absent signal in Western blot despite expecting AVPI1 expression:

• Protein extraction optimization:

  • Use lysis buffers containing appropriate detergents (RIPA, NP-40)

  • Include protease inhibitor cocktail

  • Maintain cold temperature throughout extraction

• Technical adjustments:

  • Protein loading: Increase loaded protein amount (start with 25-50 μg)

  • Transfer parameters: Optimize for low molecular weight proteins (~21.5 kDa)

  • Membrane type: PVDF may provide better retention for certain antibodies

  • Blocking agent: Try alternatives (BSA vs. milk) if background is problematic

  • Primary antibody incubation: Extend to overnight at 4°C

  • Detection system: Consider more sensitive chemiluminescence reagents

• Controls:

  • Include lysate from cells transfected with AVPI1 expression vector as positive control

  • Use β-actin or GAPDH as loading controls

What are the optimal storage conditions for AVPI1 antibodies to maintain activity?

Based on manufacturer recommendations:

• Storage temperature: -20°C (not -80°C)

• Storage buffer: PBS with 0.02% sodium azide and 50% glycerol at pH 7.3

• Stability: Typically stable for one year after shipment when properly stored

• Aliquoting: For most preparations, aliquoting is unnecessary for -20°C storage

• Freeze-thaw cycles: Minimize repeated freeze-thaw cycles

How can I optimize dual immunofluorescence with AVPI1 and other markers?

For co-localization studies using AVPI1 antibodies with other markers:

• Primary antibody selection:

  • Choose AVPI1 antibodies raised in different host species than your other target antibodies

  • If using multiple rabbit antibodies, consider directly conjugated antibodies or sequential immunostaining protocols

• Protocol optimization:

  • Titrate each primary antibody individually before combining

  • Test different fixation methods to preserve both antigens

  • Optimize antigen retrieval conditions compatible with both targets

  • Use appropriate controls to confirm specificity of each signal

• Cross-reactivity prevention:

  • Include additional blocking steps between sequential antibody applications

  • Consider using fragment antibodies (Fab) to prevent cross-reactivity

  • Use highly cross-adsorbed secondary antibodies

How can I study the relationship between AVPI1 and epithelial sodium channel (ENaC) regulation?

Given AVPI1's potential role in ENaC down-regulation , these methodological approaches would be valuable:

• Expression correlation studies:

  • Measure AVPI1 and ENaC subunit expression levels in the same tissues/cells

  • Use qPCR, Western blot, and immunofluorescence for comprehensive analysis

• Functional assays:

  • Electrophysiological measurements of ENaC activity after AVPI1 modulation

  • Amiloride-sensitive current recordings in cells with AVPI1 knockdown or overexpression

  • Surface biotinylation assays to measure ENaC membrane expression

• Interaction studies:

  • Co-immunoprecipitation of AVPI1 with ENaC subunits

  • Proximity ligation assays to detect potential interactions in situ

  • GST pull-down assays with recombinant proteins

What considerations are important when using AVPI1 antibodies in neuroscience research?

The search results indicate connections between arginine vasopressin receptor systems and neurological functions , suggesting potential relevance for AVPI1 studies in neuroscience:

• Experimental design considerations:

  • Include appropriate neuronal cell types or brain regions where vasopressin signaling is active

  • Consider species differences in expression patterns, particularly between rodents and primates

  • Use transgenic models when available to understand functional significance

• Technical recommendations:

  • For brain tissue IHC, optimize fixation time carefully to preserve antigenicity

  • Consider perfusion fixation for optimal preservation of neural tissue

  • When examining neuronal cultures, co-stain with neuronal markers to confirm cell type-specific expression

• Validation approaches:

  • Include positive control tissues with known AVPI1 expression

  • Compare expression patterns with related proteins in the vasopressin signaling pathway

How should I approach studying AVPI1 in visceral hypersensitivity research?

Based on research showing involvement of the related arginine-vasopressin receptor 1a (Avpr1a) in visceral hypersensitivity , researchers studying AVPI1 in this context should consider:

• Tissue selection:

  • Focus on distal colon tissue, which has shown differential gene expression related to vasopressin signaling

  • Compare expression between hypersensitivity models and controls

• Methodological approaches:

  • Correlate AVPI1 expression with pain ratings or behavioral measures

  • Examine co-expression with Avpr1a in relevant tissues

  • Consider neuronal hyperresponsiveness assessments in the enteric nervous system

• Translational relevance:

  • Compare findings between preclinical models and human patient samples

  • Investigate potential as a therapeutic target for visceral pain conditions