spdl1 Antibody

What is SPDL1 and what cellular functions does it regulate?

SPDL1 (Spindle Apparatus Coiled-Coil Protein 1) is a coiled-coil domain containing protein that plays crucial roles in ensuring faithful mitosis and maintaining DNA fidelity. It is fundamentally involved in spindle pole body duplication and centrosome maturation, processes essential for proper cell division and genomic stability maintenance . SPDL1 functions as an important determinant of genomic stability in cells, with its dysregulation potentially contributing to chromosomal instability, which is a hallmark feature in various cancers including colorectal cancer . Recent research has also implicated SPDL1 in cell migration processes, though its effects appear to be tissue-specific, as demonstrated by contrasting findings in colorectal cancer versus osteosarcoma models .

What types of SPDL1 antibodies are available for research applications?

Commercially available SPDL1 antibodies include polyclonal antibodies such as the PACO20599, which is derived from rabbits and specifically designed for detecting human SPDL1 protein . These antibodies are typically validated for applications including Western blotting, ELISA, and immunohistochemistry (IHC) . When selecting an antibody, researchers should consider the host species (commonly rabbit), the applications for which it has been validated, and the specific epitope it recognizes. The selection should align with the experimental design and the cellular compartment being investigated, as SPDL1 expression can be predominantly cytoplasmic, with some tissues showing combined cytoplasmic-membranous (9.30%) or cytoplasmic-nuclear (6.98%) expression patterns .

What is the typical subcellular localization pattern of SPDL1 in normal versus cancer tissues?

In colorectal cancer tissues, SPDL1 expression is predominantly observed in the cytoplasm, characterized by ubiquitous, diffuse staining on tumor cells though not with uniform intensity. Some cancer tissues exhibit combined expression patterns:

• Cytoplasmic-membranous: 9.30% of cases

• Cytoplasmic-nuclear: 6.98% of cases

Notably, only a small subset of cells within tumor glands display nuclear staining patterns. In contrast, non-cancerous adjacent tissues show staining exclusively restricted to the cytoplasm . This differential localization pattern can serve as an important distinguishing feature when analyzing SPDL1 expression in normal versus malignant tissues and may reflect altered protein function in the cancer context.

What are the validated applications for SPDL1 antibodies and their recommended protocols?

According to available product information, SPDL1 antibodies have been validated for the following applications with specific recommended dilutions:

When using these antibodies for IHC analysis, researchers should note that SPDL1 shows predominantly cytoplasmic staining in both normal and cancer tissues, with some variation in expression patterns . For optimal results, researchers should perform antibody titration experiments to determine the ideal concentration for their specific tissue type and fixation method.

How should researchers quantify and interpret SPDL1 immunohistochemical staining in tissue samples?

For accurate quantification of SPDL1 expression in tissue sections, researchers should employ a systematic scoring approach. Based on published research methodologies, intensity scoring can be categorized as:

• Low expression

• Moderate expression

• High expression

When establishing cut-off points for expression categories, researchers may utilize statistical methods such as receiver operating characteristic (ROC) curve analysis . In survival studies, it's important to note that moderate expression levels of SPDL1 have been associated with particularly poor prognosis in colorectal cancer, while high expression levels correlate with improved prognosis .

For accurate interpretation, researchers should consider:

• The predominant cellular localization (cytoplasmic, nuclear, or membranous)

• The percentage of positive cells

• The staining intensity

• The pattern (diffuse vs. focal)

Additionally, comparison with adjacent normal tissue is essential for contextualizing expression levels, as research has shown significantly higher SPDL1 expression in colorectal cancer tissues compared to adjacent non-cancerous tissues (p = 0.004) .

How does SPDL1 expression correlate with clinicopathological features and patient outcomes in cancer research?

Research on colorectal cancer has provided valuable insights into SPDL1's clinical significance. High SPDL1 expression in tumors has been associated with improved survival outcomes compared to low expression counterparts, even after adjustment for multiple confounding factors. Specifically:

• Age

• Gender

• Tumor grading

• Primary tumor (pT) classification

• Regional lymph node (pN) status

• Distant metastasis (pM) status

• AJCC TNM stage

• Vascular invasion

• Perineural invasion

• Resection margins

• Primary tumor location

This suggests that SPDL1 may function as an independent biomarker rather than being associated with specific tumor characteristics.

How does SPDL1 relate to genomic instability markers in cancer research?

SPDL1 expression shows significant associations with various genomic instability phenotypes and markers:

• Higher expression in chromosomal instability (CIN) subtype tumors compared to genomically stable (GS) tumors (p < 0.0001)

• Higher expression in microsatellite instability (MSI) tumors compared to GS tumors (p = 0.002)

• Positive correlation with CIN (r = 0.30; p < 0.0001) and MSI (r = 0.33; p = 0.002)

• Association with aneuploidy score (r = 0.13; p = 0.03)

• Strong correlation with proliferation marker MKI67 (r = 0.56; p < 0.0001)

• Significant correlation with DNA mismatch repair genes MSH6 (r = 0.58), MSH2 (r = 0.58), MLH1 (r = 0.35), and PMS2 (r = 0.35)

Additionally, SPDL1 shows correlation with almost all 25 genes associated with functional aneuploidy (the CIN25 signature), including FOXM1 (r = 0.53; p < 0.0001), though notably not with TP53 . These associations suggest SPDL1's integral role in genomic stability mechanisms and highlight its potential utility as a biomarker for specific molecular subtypes of cancer.

How should researchers address tissue-specific variations in SPDL1 function and prognostic significance?

The prognostic significance of SPDL1 appears to be tissue-specific, with contrasting findings across different cancer types. For instance:

• In colorectal and pancreatic ductal adenocarcinoma: SPDL1 overexpression correlates with better outcomes

• In oral squamous cell carcinoma: SPDL1 overexpression relates to chromosomal instability and worse outcomes

When designing studies to assess SPDL1's role in specific cancer types, researchers should:

• Include tissue-specific controls and compare with normal adjacent tissue

• Consider potential stage-specific effects, as SPDL1's impact may vary by disease stage

• Analyze SPDL1 in the context of the tumor's genetic background and chromosomal instability status

• Examine SPDL1's relationship with tissue-specific cell migration patterns, as its effects on invasion and migration appear to differ between cancer types

• Employ multiple methodologies (IHC, gene expression analysis) to comprehensively assess SPDL1's role

This approach acknowledges that SPDL1 functions as a "double-faced protein" whose levels need tight regulation in tumors, with moderate expression potentially conferring different outcomes than high expression .

What are the key considerations for designing experiments to study SPDL1's role in cell migration and invasion?

When investigating SPDL1's role in cell migration and invasion, researchers should note the contradictory findings in different cancer models. In designing robust experiments:

• Cell line selection: Include multiple cell lines from the same cancer type to account for heterogeneity

• Experimental approaches: Combine 2D and 3D migration assays, as SPDL1-depleted primary fibroblasts and U2OS cells migrated slower in 2D culture, while the opposite effect was observed in other cancer models

• SPDL1 modulation techniques:

  • siRNA knockdown

  • CRISPR-Cas9 knockout

  • Overexpression models using expression vectors

• Downstream analysis:

  • Assess cytoskeletal changes using immunofluorescence

  • Examine proteins involved in cell adhesion and invasion

  • Evaluate changes in related signaling pathways

• In vivo validation: Consider using xenograft models to confirm in vitro findings

These experiments should include appropriate controls and be designed to detect both promotion and inhibition of migration/invasion, accounting for the context-dependent role of SPDL1 observed across different cancer types .

How can researchers differentiate between SPDL1's roles in chromosome instability versus cell migration?

SPDL1's dual functions in chromosome stability and cell migration present a complex experimental challenge. To differentiate between these roles, researchers should design a strategic experimental approach:

• Temporal analysis: Monitor SPDL1 localization throughout the cell cycle using time-lapse microscopy with fluorescently-tagged SPDL1

• Domain-specific mutations: Create constructs with mutations in specific SPDL1 domains to selectively disrupt either its spindle-associated functions or migration-related interactions

• Pathway inhibition studies: Use specific inhibitors of mitotic processes versus migration signaling pathways to determine which SPDL1-associated function is affected

• Chromosome stability assessments:

  • Analyze aneuploidy via karyotyping or FISH

  • Measure mitotic errors using live cell imaging

  • Quantify γ-H2AX foci formation as a marker of DNA damage

• Migration-specific readouts:

  • Wound healing assays

  • Transwell migration/invasion assays

  • Real-time analysis of cell motility parameters

By systematically comparing these readouts in SPDL1-modulated versus control cells, researchers can decouple the protein's distinct functions and determine whether its prognostic significance in different cancers stems primarily from one function or represents a combined effect of both roles .

How might SPDL1 antibodies be integrated into immune checkpoint blockade research?

While the search results primarily focus on SPDL1 rather than immune checkpoint proteins, there may be interesting connections to explore between SPDL1 and immune checkpoint molecules. Given that search result #2 mentions sPD1 and sPDL1 as biomarkers for evaluating immune states in cancer patients , researchers might consider:

• Co-expression studies: Investigate potential correlations between SPDL1 expression and immune checkpoint molecules like PD1/PDL1 in tumor samples

• Immune cell profiling: Examine whether SPDL1 expression in tumors correlates with tumor-infiltrating lymphocyte characteristics or specific immune cell populations

• Response prediction: Assess whether SPDL1 expression levels might complement immune checkpoint expression in predicting response to immunotherapy

• Combination markers: Develop multiplexed immunohistochemistry protocols using antibodies against both SPDL1 and immune checkpoint molecules to create more comprehensive prognostic profiles

While direct evidence linking SPDL1 to immune checkpoint pathways is currently limited, this represents an intriguing avenue for future research, particularly given the importance of both genomic instability and immune evasion as hallmarks of cancer.

What are the current technical limitations of SPDL1 antibodies and potential directions for improvement?

Based on the available information, researchers working with SPDL1 antibodies should be aware of several technical considerations:

• Specificity verification: Current antibodies should be validated using positive and negative controls, including SPDL1 knockdown/knockout samples, to ensure specificity

• Isoform detection: It's unclear whether current antibodies detect all potential SPDL1 isoforms, which may be important for comprehensive analysis

• Cross-reactivity: Researchers should verify species cross-reactivity, as the described antibody is validated for human samples but may have limited utility in model organisms

• Application scope: While validated for ELISA and IHC, expansion to other applications like immunofluorescence, flow cytometry, and ChIP might enhance research capabilities

Future directions for antibody development might include:

• Generating isoform-specific antibodies

• Developing antibodies with broader species cross-reactivity

• Creating phospho-specific antibodies to detect activated forms of SPDL1

• Producing high-affinity monoclonal antibodies with defined epitopes for more consistent results across studies

These advancements would enable more detailed investigations into SPDL1's complex roles in normal and cancer biology.