ndel1b Antibody

Introduction to NDEL1b Protein and Its Antibodies

NDEL1b is one of two distinct NDEL1 orthologues (Ndel1a and Ndel1b) found specifically in zebrafish (Danio rerio), resulting from a gene duplication event that occurred during teleost evolution . Unlike mammals, which possess only the paralogous genes NDE1 and NDEL1, zebrafish have undergone an additional duplication of NDEL1, creating a more complex gene family with potentially specialized functions . NDEL1b antibodies are immunoglobulins specifically designed to target and bind to the NDEL1b protein, enabling its detection, quantification, and characterization in research settings.

The evolution of NDEL1 proteins follows a fascinating trajectory, with NDEL1b representing a specialized adaptation in zebrafish:

Production and Validation of NDEL1b Antibodies

NDEL1b antibodies are typically produced through either polyclonal or monoclonal methods, each with distinct advantages:

Validation of NDEL1b antibodies typically employs multiple techniques to ensure specificity, including:

1. Western blotting to confirm the detection of appropriately sized bands specific to NDEL1b

2. Immunoprecipitation to verify the ability to isolate NDEL1b from complex protein mixtures

3. Immunohistochemistry or immunofluorescence to confirm expected tissue localization patterns

4. Peptide competition assays to demonstrate binding specificity

5. Knockout/knockdown controls to verify absence of signal when the target is removed

Applications of NDEL1b Antibodies in Research

NDEL1b antibodies serve as valuable tools across multiple research applications, particularly in zebrafish developmental neuroscience:

Western Blotting (WB)

NDEL1b antibodies are commonly used in Western blotting applications, with recommended dilutions typically ranging from 1:500 to 1:2000 . In zebrafish samples, NDEL1b typically appears as a band at approximately 38-42 kDa on SDS-PAGE gels, consistent with its calculated molecular weight of approximately 38 kDa . Western blotting applications allow researchers to quantify NDEL1b expression changes during developmental processes or experimental interventions.

Immunohistochemistry (IHC)

In fixed zebrafish tissue sections, NDEL1b antibodies enable visualization of protein localization patterns. Recommended dilutions for IHC applications typically range from 1:50 to 1:200 . The predominantly neuronal expression pattern of NDEL1 family proteins makes IHC particularly valuable for studying neurodevelopmental processes in zebrafish models.

Immunofluorescence (IF)

NDEL1b antibodies conjugated to fluorophores or used with secondary fluorescent antibodies allow for high-resolution imaging of NDEL1b localization within cells. This application has revealed that, like its mammalian counterpart, NDEL1b likely localizes to the centrosome and mitotic spindle during cell division .

Immunoprecipitation (IP)

NDEL1b antibodies can be used to isolate NDEL1b protein complexes from zebrafish samples, enabling the identification of interaction partners through subsequent mass spectrometry analysis . This approach has proven valuable for understanding the protein interaction networks involving NDEL1 family proteins.

NDEL1b in Zebrafish Neurodevelopment

The zebrafish model offers unique advantages for studying neurodevelopmental processes due to its external development, optical transparency, and genetic tractability. NDEL1b antibodies have facilitated research revealing that:

1. NDEL1b is expressed in developing neural tissues of zebrafish embryos

2. NDEL1b likely plays crucial roles in neuronal migration similar to mammalian NDEL1

3. The expression patterns of NDEL1b may differ from NDEL1a, suggesting functional specialization

The duplication of NDEL1 in zebrafish into NDEL1a and NDEL1b suggests an evolutionary advantage through subfunctionalization or neofunctionalization, making the zebrafish model particularly valuable for understanding the ancestral and derived functions of this gene family.

Interaction Partners of NDEL1b

NDEL1b antibodies have helped identify key interaction partners of the NDEL1b protein, providing insights into its cellular functions:

Like mammalian NDEL1, zebrafish NDEL1b appears capable of forming homodimers and homotetramers, as well as potential heteromers with NDEL1a and NDE1 . These oligomeric states likely play important roles in regulating protein function and interactions.

Technical Considerations for NDEL1b Antibody Usage

When working with NDEL1b antibodies, several technical considerations can optimize results:

1. Storage: Most NDEL1b antibodies should be stored at -20°C for long-term stability, with short-term storage at 4°C for up to one month . Repeated freeze-thaw cycles should be avoided.

2. Working Dilutions: Optimal dilutions vary by application:

   • Western Blot: 1:500-1:2000

   • Immunohistochemistry: 1:50-1:200

   • Immunofluorescence: 1:100-1:500

3. Controls: Appropriate positive controls (zebrafish brain tissue) and negative controls (pre-immune serum or isotype-matched control antibodies) should be included in experimental designs.

4. Cross-reactivity: Due to the high homology between NDEL1a and NDEL1b in zebrafish, thorough validation is required to ensure selective detection of NDEL1b.

Future Perspectives in NDEL1b Antibody Research

The continued development and application of NDEL1b antibodies promise several advances:

1. Creation of highly specific monoclonal antibodies capable of distinguishing between NDEL1a and NDEL1b in zebrafish

2. Development of phospho-specific NDEL1b antibodies to study regulatory post-translational modifications

3. Application in high-throughput screening approaches to identify compounds affecting NDEL1b function

4. Integration with emerging proteomics technologies to comprehensively map NDEL1b interaction networks

5. Utilization in comparative studies between zebrafish and mammalian models to understand evolutionarily conserved and divergent functions