GPD1L Antibody

What is GPD1L and what is its physiological function?

GPD1L is encoded by the gene GPD1L, which is located on chromosome 3p22.3. It functions primarily as an enzyme responsible for catalyzing the conversion of sn-glycerol 3-phosphate to glycerone phosphate. The protein has a calculated molecular weight of 351 amino acids (38 kDa), though the observed molecular weight in Western blot applications typically ranges from 35-40 kDa . This enzyme plays important roles in cellular metabolism and has been found to be dysregulated across various cancer types, suggesting its potential importance in both normal physiology and pathological states .

Which applications are GPD1L antibodies validated for?

Based on current research, GPD1L antibodies have been primarily validated for Western Blot (WB) and ELISA applications. Different antibody formulations show varying recommended dilutions - for instance, antibody 17263-1-AP is recommended at dilutions of 1:500-1:2000 for Western Blot applications, while antibody 83792-2-RR requires higher dilutions ranging from 1:5000-1:50000 . It is important to note that these antibodies should be titrated in each specific testing system to obtain optimal results, as performance can be sample-dependent .

What positive controls are recommended for GPD1L antibody validation?

For robust experimental design, researchers should consider using the following validated positive controls:

These validated controls provide essential reference points when optimizing antibody-based detection methods and ensuring specificity in experimental protocols .

What are the optimal storage conditions for maintaining GPD1L antibody activity?

To maintain optimal activity, GPD1L antibodies should be stored at -20°C, where they remain stable for approximately one year after shipment. The antibodies are typically supplied in a storage buffer consisting of PBS with 0.02% sodium azide and 50% glycerol at pH 7.3. Importantly, for -20°C storage, aliquoting is considered unnecessary according to manufacturer recommendations, though this may depend on the specific antibody formulation and frequency of use. Some smaller volume preparations (20μl sizes) may contain 0.1% BSA as a stabilizer .

How should Western blot protocols be optimized for GPD1L detection?

For optimal GPD1L detection via Western blot, researchers should consider the following methodological recommendations:

• Use the appropriate antibody dilution for your specific antibody (1:500-1:2000 for 17263-1-AP; 1:5000-1:50000 for 83792-2-RR)

• Ensure adequate sample preparation that preserves protein integrity

• Include both positive controls (e.g., heart tissue, HEK-293 cells) and negative controls

• Target the correct molecular weight range (35-40 kDa) for band identification

• Consider antigen retrieval methods if necessary for your specific sample type

• Follow manufacturer-specific protocols for optimal results

What cross-reactivity considerations are important when selecting a GPD1L antibody?

GPD1L antibodies have demonstrated reactivity across human, mouse, and rat samples in validated studies. When selecting an antibody for experimental use, researchers should consider whether their target species has been validated. For antibodies like 17263-1-AP and 83792-2-RR, cross-reactivity with human, mouse, and rat has been established, making them suitable choices for experiments involving these species. For other species, additional validation would be required. The immunogen information (GPD1L fusion protein Ag10977) can provide insight into potential cross-reactivity patterns .

How can siRNA knockdown experiments be validated using GPD1L antibodies?

For validating GPD1L knockdown experiments, a systematic approach using antibody-based detection is essential:

• Perform Western blot analysis with validated GPD1L antibodies to confirm protein reduction

• Include appropriate positive controls (cells/tissues known to express GPD1L) for comparison

• Quantify knockdown efficiency through densitometry analysis

• Consider including multiple time points post-transfection to assess knockdown stability

• Validate knockdown effects through functional assays

Research has demonstrated the utility of this approach, as in one study where GPD1L knockdown in Hep3B cells via siRNA was confirmed using antibody detection, revealing altered sensitivity to the therapeutic agent PF-562271 .

How do different fixation and sample preparation methods affect GPD1L detection?

While specific comparative data on fixation methods is not provided in the search results, general considerations for GPD1L detection include:

• For Western blot applications, protein extraction methods should preserve native protein structure

• Different lysis buffers may yield varying results depending on GPD1L's subcellular localization

• For immunohistochemistry, fixation methods (paraformaldehyde vs. formalin) may affect epitope accessibility

• Fresh-frozen vs. FFPE (formalin-fixed paraffin-embedded) samples may require different antibody concentrations

• Antigen retrieval methods may be necessary for some sample types

Researchers should conduct preliminary optimization experiments to determine the ideal sample preparation method for their specific experimental context.

What are the contrasting roles of GPD1L in different cancer types?

Research has revealed intriguing and contrasting roles for GPD1L across different cancer types:

• In hepatocellular carcinoma (HCC):

  • High GPD1L expression correlates with poor patient survival

  • Expression increases with advancing tumor stage, suggesting positive selection during tumorigenesis

  • Associated with adverse histological characteristics including vascular invasion and higher histological grade

  • Spatial and single-cell transcriptome datasets confirm elevated GPD1L expression in tumor tissue compared to adjacent normal tissue

• In pancreatic ductal adenocarcinoma (PDAC):

  • GPD1L is down-regulated

  • Down-regulation is associated with poorer prognosis

  • GPD1L is targeted by a three-microRNA signature (miR155, miR181a, and miR221)

These contrasting expression patterns underscore the tissue-specific and context-dependent role of GPD1L in cancer biology, highlighting the importance of careful experimental design when studying its function in different cancer types.

How can GPD1L expression serve as a predictive biomarker for therapeutic response?

Analysis of GPD1L as a potential predictive biomarker has yielded promising results, particularly in HCC:

• GPD1L mRNA levels in 17 HCC cell lines showed a robust inverse correlation with therapeutic response (measured by IC50) for three agents:

  • PF-562271 (a focal adhesion kinase inhibitor)

  • Linsitinib

  • BMS-754807

• In vitro validation demonstrated that cell lines with lower GPD1L expression (PLC/PRF/5 and HepG2) exhibited greater resistance to these drugs compared to a cell line with higher GPD1L expression (Hep3B)

• Knockdown of GPD1L in Hep3B cells specifically reduced sensitivity to PF-562271, while sensitivity to the other compounds remained unchanged

These findings suggest GPD1L expression levels could potentially guide therapeutic selection in HCC patients, though additional clinical validation is needed.

What is the relationship between GPD1L and microRNA regulation in cancer?

In pancreatic ductal adenocarcinoma (PDAC), research has identified a regulatory relationship between GPD1L and microRNAs:

• A three-microRNA signature (miR155, miR181a, and miR221) was found to be upregulated in PDAC

• Bioinformatic analysis identified GPD1L as having potential binding sites for all three microRNAs

• Experimental validation confirmed that GPD1L is down-regulated in PDAC samples

• This down-regulation correlates with poorer prognosis across three independent datasets

This regulatory mechanism provides insight into how GPD1L expression is controlled in cancer and suggests potential therapeutic strategies targeting these microRNA-GPD1L interactions.

How can single-cell and spatial transcriptomics data be integrated with GPD1L antibody-based protein detection?

Modern research increasingly combines multi-omics approaches for comprehensive understanding:

• Validation of transcriptomic findings at the protein level through immunohistochemistry with GPD1L antibodies

• Correlation of single-cell RNA expression patterns with protein expression in serial tissue sections

• Multiplexed immunofluorescence to co-localize GPD1L with other markers of interest

• Integration of spatial transcriptomics data to map GPD1L expression within heterogeneous tumor tissues

• Computational approaches to integrate protein and RNA data across spatial dimensions

Research on HCC has already begun implementing such approaches, with spatial and single-cell transcriptome datasets confirming elevated GPD1L expression in tumor tissue compared to adjacent normal tissue .

What considerations are important when comparing results from different GPD1L antibodies?

When comparing experimental results obtained using different GPD1L antibodies, researchers should consider:

• Epitope differences: Antibodies may target different regions of GPD1L, affecting detection efficiency

• Antibody format: Polyclonal (like 17263-1-AP) versus recombinant (like 83792-2-RR) antibodies have different properties

• Purification methods: Antigen affinity purification versus Protein A purification may affect specificity

• Recommended dilutions: These vary significantly between antibodies (1:500-1:2000 versus 1:5000-1:50000)

• Validation status: Consider the extent of validation in published literature and manufacturer data

These differences highlight the importance of antibody validation and consistent methodology when comparing results obtained with different GPD1L antibodies.