PDE1B Antibody, Biotin conjugated

What is PDE1B and why is it a significant research target?

PDE1B (Phosphodiesterase 1B) is a calcium/calmodulin-dependent 3',5'-cyclic nucleotide phosphodiesterase that demonstrates 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 through its enzymatic activity . PDE1B is primarily localized in the cytoplasm and functions as a homodimer .

The significance of PDE1B as a research target stems from its involvement in critical cellular signaling pathways. Recent studies have demonstrated that PDE1B expression increases under pathological conditions such as ischemia, where it has been shown to progressively elevate in the peri-infarct region after focal middle cerebral artery occlusion . This makes PDE1B an important target for neurological research, particularly in understanding microglial activation and neuroinflammation.

What are the key specifications of commercially available PDE1B Antibody, Biotin conjugated?

PDE1B Antibody, Biotin conjugated is typically available as a polyclonal antibody raised in rabbits . The antibody is targeted against specific peptide sequences of the PDE1B protein, with some products using synthetic peptides corresponding to unique amino acid sequences on the PDE1B gene , while others utilize recombinant human calcium/calmodulin-dependent 3',5'-cyclic nucleotide phosphodiesterase 1B protein (specifically amino acids 1-277) .

The antibody is supplied in liquid form with a concentration of approximately 0.579 μg/μl in antibody stabilization buffer . Storage buffers may contain preservatives such as 0.03% Proclin 300 and constituents like 50% Glycerol in 0.01M PBS at pH 7.4 . The recommended storage condition is -20°C for long-term storage, with caution advised against repeated freeze-thaw cycles .

What experimental applications is PDE1B Antibody, Biotin conjugated suitable for?

PDE1B Antibody, Biotin conjugated has been validated for multiple experimental applications:

• Chromatin immunoprecipitation (CM): 1:50-1:200 dilution

• Enzyme-linked immunosorbent assay (ELISA): 1:10,000 dilution

• Immunocytochemistry (ICC): 1:50-1:200 dilution

• Immunofluorescence (IF): 1:50-1:200 dilution

• Immunohistochemistry (IHC): 1:50-1:200 dilution

• Immunoprecipitation (IP): 1:200 dilution

• Western blotting (WB): 1:500-1:1,000 dilution

These applications make the antibody versatile for examining PDE1B expression, localization, and interactions across various experimental systems. The biotin conjugation is particularly advantageous for detection systems utilizing streptavidin, enhancing signal amplification and detection sensitivity.

What is the specificity profile of PDE1B Antibody, Biotin conjugated?

The specificity of PDE1B Antibody, Biotin conjugated is a critical consideration for experimental design. High-quality antibodies do not cross-react with other PDE family members, including the closely related PDE1A or PDE1C isoforms . The antibody is designed to label all PDE1B variants, providing comprehensive detection of the target protein .

When validating antibody specificity in your experimental system, it is recommended to include appropriate positive and negative controls. For negative controls, samples known not to express PDE1B or knockdown/knockout systems can be used. The specificity can be further verified through western blotting, where PDE1B appears as a band at approximately 63 kDa (the expected molecular weight for Calcium/calmodulin-dependent 3',5'-cyclic nucleotide phosphodiesterase 1B) .

What species reactivity does PDE1B Antibody, Biotin conjugated demonstrate?

PDE1B Antibody, Biotin conjugated products show varying species reactivity profiles depending on the manufacturer. Some antibodies are reactive with human, mouse, and rat samples , while others may be specifically optimized for human samples . This species reactivity information is crucial for selecting the appropriate antibody for your experimental model.

The cross-species reactivity is determined by the conservation of the epitope sequence across species. When working with species not listed in the manufacturer's specifications, additional validation steps should be undertaken to confirm reactivity. This may include positive control tissues from the species of interest and sequence homology analysis of the immunogen region.

How can PDE1B inhibition affect microglial exosome release and neuronal protection?

Recent research has revealed a fascinating role for PDE1B in the regulation of microglial function and exosome release under ischemic conditions. Inhibition of PDE1B by vinpocetine in microglial cells has been shown to promote the M2 anti-inflammatory phenotype while inhibiting the pro-inflammatory M1 phenotype . This modulation of microglial polarization has significant implications for neuroinflammatory responses.

Mechanistically, knockdown or inhibition of PDE1B significantly enhances autophagic flux in BV2 microglial cells, with vinpocetine-mediated suppression of the M1 phenotype being dependent on autophagy under ischemic conditions . The experimental approach involves:

• Establishment of oxygen-glucose-deprivation (OGD) models to simulate ischemic conditions

• Treatment with PDE1B inhibitors such as vinpocetine

• Assessment of microglial polarization markers (CD11b for M1 and Arg-1 for M2)

• Evaluation of autophagic flux through LC3-II expression

Furthermore, PDE1B inhibition alters exosome biogenesis in OGD-treated microglial cells. Exosomes released from vinpocetine-treated OGD-BV2 cells have been shown to reverse the neuronal cell apoptosis and neurite dysfunction induced by exosomes from untreated OGD-BV2 cells . This protective effect highlights the potential therapeutic implications of targeting PDE1B in ischemic brain injury.

What methodological approaches can optimize the performance of PDE1B Antibody, Biotin conjugated in experimental settings?

To achieve optimal performance with PDE1B Antibody, Biotin conjugated, several methodological considerations should be addressed:

How does PDE1B expression change under pathological conditions, and what are the implications for experimental design?

PDE1B expression demonstrates dynamic changes under pathological conditions, particularly in ischemic brain injury models. In the peri-infarct region following middle cerebral artery occlusion (MCAO), Iba-1-positive microglial cells significantly increase on the first day and peak at the 14th day post-MCAO . Concurrently, PDE1B expression is progressively elevated in these regions, correlating with microglial activation.

In vitro studies using oxygen-glucose deprivation (OGD) models confirm that PDE1B expression increases in OGD-treated microglial BV2 cells . This upregulation coincides with changes in microglial polarization markers, with increased CD11b (M1 marker) and decreased Arg-1 (M2 marker) expression.

These expression patterns have important implications for experimental design:

• Temporal considerations: When investigating PDE1B in pathological conditions, time-course experiments are crucial to capture the dynamic expression changes.

• Marker co-localization: Co-staining analysis should be employed to determine PDE1B expression in specific cell populations, such as CD11b-positive or Arg-1-positive microglia.

• Model selection: Researchers should note the differences between in vivo and in vitro models. For example, while Iba-1 expression increases in activated microglia in vivo, it may decrease in BV2 cells under OGD conditions in vitro .

• Inhibitor studies: When using PDE1B inhibitors like vinpocetine, dose-dependent effects should be characterized to establish optimal experimental conditions.

What are the technical considerations for using PDE1B Antibody, Biotin conjugated in studying autophagy-related pathways?

PDE1B has emerging roles in autophagy regulation, particularly in microglial cells under ischemic conditions. When designing experiments to investigate these pathways using PDE1B Antibody, Biotin conjugated, several technical considerations should be addressed:

• Autophagic flux assessment: PDE1B inhibition enhances autophagic flux in BV2 cells, as evidenced by increased LC3-II levels . Proper assessment of autophagic flux requires:

  • Comparison of LC3-II levels with and without lysosomal inhibitors (e.g., bafilomycin A1)

  • Monitoring p62/SQSTM1 degradation

  • Evaluating autophagosome-lysosome fusion

• Dual fluorescence detection: When using biotin-conjugated PDE1B antibody in immunofluorescence studies of autophagy, optimize protocols for simultaneous detection of:

  • PDE1B (using streptavidin-conjugated fluorophores)

  • Autophagy markers (LC3, p62, LAMP1, etc.)

  • Cell type-specific markers (Iba-1 for microglia)

• Pharmacological manipulations: Include appropriate controls when combining PDE1B antibody detection with autophagy modulators:

  • PDE1B inhibitors (vinpocetine)

  • Autophagy inhibitors (3-Methyladenine)

  • Autophagy inducers (rapamycin)

• siRNA validation: When using siRNA to knockdown PDE1B expression, validate knockdown efficiency through western blotting and correlate with changes in autophagic flux .

These methodological approaches enable robust investigation of the relationship between PDE1B activity and autophagy regulation, particularly in the context of neuroinflammatory responses.