PDE1B Antibody

What is PDE1B and why is it important to study?

PDE1B (Phosphodiesterase 1B) is a calcium/calmodulin-dependent cyclic nucleotide phosphodiesterase that exhibits dual specificity for the second messengers cAMP and cGMP, with a preference for cGMP as a substrate. It functions as a key regulator of many important physiological processes . PDE1B belongs to the PDE1 subfamily and is stimulated by calcium-calmodulin complex. It plays crucial roles in signal transduction pathways, particularly in neuronal tissues and has been implicated in various disease processes including cancer .

When selecting a PDE1B antibody, researchers should consider:

• Antibody Type: Monoclonal antibodies offer high specificity for particular epitopes, while polyclonal antibodies provide broader detection capability .

• Species Reactivity: Ensure the antibody reacts with your species of interest. Available PDE1B antibodies have been validated for reactivity with human, mouse, rat, monkey, and dog samples .

• Binding Region: Different antibodies target specific regions of PDE1B:

  • AA 370-536 region antibodies

  • AA 473-500 (C-Terminal) antibodies

  • AA 1-277 region antibodies

  • Full-length protein antibodies

• Validated Applications: Confirm the antibody has been validated for your intended application. For example, if performing IHC-P, verify the antibody has been tested for this specific application .

• Detection Method: Consider whether unconjugated antibodies or those with specific conjugates are needed based on your detection system .

What are the optimal protocols for Western blotting with PDE1B antibodies?

For optimal Western blotting results with PDE1B antibodies:

• Sample Preparation:

  • Use brain tissue (particularly cerebellum or cerebrum) as positive controls

  • Load approximately 20 μg of protein lysate per lane

• Antibody Dilution:

  • Primary antibody: 1:1000-1:4000 is commonly used for most PDE1B antibodies

  • Secondary antibody: typically 1:2000 for HRP-conjugated antibodies

• Incubation Conditions:

  • Room temperature incubation for 1.5 hours has been validated for certain PDE1B antibodies

• Expected Results:

  • Predicted band size: 61 kDa

  • Observed band size: 59-61 kDa (may vary by sample type)

Published Western blot data shows clear bands at approximately 61 kDa in human cerebellum and brain lysates, confirming antibody specificity .

How should immunohistochemistry be performed with PDE1B antibodies?

For successful immunohistochemistry with PDE1B antibodies:

• Sample Preparation:

  • Use formalin/PFA-fixed paraffin-embedded sections

  • Brain tissue (cerebrum/cerebellum) serves as excellent positive control material

• Antigen Retrieval:

  • Heat-mediated antigen retrieval using Bond™ Epitope Retrieval Solution 1 (pH 6.0) has been validated

• Antibody Dilution and Detection:

  • Dilution ranges from 1:50-1:2000 depending on the specific antibody

  • For some antibodies (e.g., ab182565), a 1:2000 dilution (0.47 μg/ml) has been validated

  • Use appropriate secondary detection systems (e.g., Rabbit specific IHC polymer detection kit HRP/DAB)

• Controls:

  • Use PBS instead of primary antibody as negative control

  • Counterstain with hematoxylin for nuclear visualization

How do I troubleshoot common issues with PDE1B antibody experiments?

Common challenges and solutions when working with PDE1B antibodies:

• Non-specific Binding:

  • Increase blocking time/concentration

  • Optimize antibody dilution (test range from 1:500-1:8000 for WB)

  • Use additional washing steps with increased detergent

• Weak Signal:

  • Confirm sample type (brain tissue shows highest expression)

  • Reduce antibody dilution

  • Increase protein loading (up to 20-30 μg)

  • Extend primary antibody incubation time

• Multiple Bands:

  • PDE1B has two isoforms: PDE1B1 (61 kDa) and PDE1B2 (59 kDa)

  • Determine which isoform your antibody detects

  • Optimize SDS-PAGE conditions for better separation

• Storage-Related Problems:

  • Aliquot and store at -20°C

  • Avoid repeated freeze/thaw cycles

  • Use storage buffer with stabilizers (PBS with 0.02% sodium azide, 50% glycerol, pH 7.3)

How can PDE1B antibodies be utilized to investigate its role in cancer biology?

Recent research has identified PDE1B as a potential tumor suppressor gene in osteosarcoma . When investigating PDE1B in cancer research:

What methodologies are recommended for studying PDE1B's role in neurological research?

PDE1B is enriched in brain tissue, making it relevant for neurological research:

• Regional Expression Mapping:

  • Use IHC with PDE1B antibodies to map expression across different brain regions

  • Western blot analysis of region-specific lysates (cerebellum, cerebrum) shows consistent expression

• Cellular Localization:

  • Employ immunofluorescence with PDE1B antibodies for subcellular localization

  • Co-staining with neuronal markers helps identify specific cell populations expressing PDE1B

• Functional Studies:

  • Use PDE1B antibodies to validate knockdown/overexpression models

  • Investigate PDE1B's role in cAMP/cGMP signaling in neuronal function

  • Study calcium-calmodulin regulation of PDE1B activity in neurons

• Disease Models:

  • Compare PDE1B expression and localization in normal vs. pathological brain tissues

  • Investigate potential alterations in neurodegenerative or psychiatric disorders

How can I validate PDE1B antibody specificity for my experimental system?

Thorough validation of PDE1B antibodies is crucial for experimental integrity:

• Positive Controls:

  • Brain tissue (cerebellum, cerebrum) is the recommended positive control

  • Human, mouse, and rat brain tissues have been validated

• Knockout/Knockdown Validation:

  • Compare antibody signal in wild-type vs. PDE1B-knockout/knockdown samples

  • Absence of signal in knockout/knockdown samples confirms specificity

• Peptide Competition:

  • Pre-incubate antibody with immunizing peptide before application

  • Signal elimination indicates antibody specificity for the target epitope

• Multiple Antibody Comparison:

  • Test antibodies targeting different PDE1B epitopes (AA 370-536, C-terminal, etc.)

  • Consistent results across different antibodies increase confidence in specificity

• Cross-Species Validation:

  • Test antibody in multiple species if appropriate for your research

  • Consistent results across species with high PDE1B homology supports specificity

How might PDE1B antibodies contribute to immunotherapy research?

Emerging evidence connects PDE1B to immune function and response to immunotherapies:

• Immune Cell Infiltration Analysis:

  • Use PDE1B antibodies in multiplex immunofluorescence to correlate expression with immune cell infiltration

  • CIBERSORT analysis has revealed associations between PDE1B expression and infiltration of 22 types of immune cells

• Tumor Microenvironment Assessment:

  • Evaluate PDE1B expression in relation to immune, stromal, and ESTIMATE scores using validated antibodies

  • Flow cytometry with PDE1B antibodies can help identify specific immune cell populations expressing PDE1B

• Immunotherapy Response Prediction:

  • Tumor Immune Dysfunction and Exclusion (TIDE) analysis suggests high PDE1B expression correlates with better immunotherapy response

  • PDE1B antibodies can help stratify patients for potential immunotherapy response

• Mechanism Investigation:

  • Explore how PDE1B prevents immune escape in tumors

  • Investigate connections to JAK-STAT and other immune-related signaling pathways

What are the considerations for studying the calcium/calmodulin regulation of PDE1B?

PDE1B is uniquely regulated by calcium/calmodulin, presenting specific experimental considerations:

• Activity Assays:

  • Use PDE1B antibodies to immunoprecipitate the enzyme for activity measurements

  • Design experiments with varying calcium concentrations to observe calmodulin-dependent activation

• Structural Analysis:

  • Different antibodies target various regions of PDE1B, allowing selective study of functional domains

  • Antibodies targeting AA 1-277 may interact with the regulatory domain while those targeting C-terminal regions interact with the catalytic domain

• Interaction Studies:

  • Use co-immunoprecipitation with PDE1B antibodies to study calmodulin binding under different calcium concentrations

  • Investigate how calmodulin binding affects PDE1B's interaction with other proteins

• Subcellular Localization Changes:

  • Employ immunofluorescence to track changes in PDE1B localization in response to calcium flux

  • Correlate with functional outcomes in signaling pathways