AGL13 Antibody

What is ALG13 and why is it important in research applications?

ALG13 (asparagine-linked glycosylation 13 homolog) is a protein involved in N-linked glycosylation pathways that has significant implications in developmental disorders. The protein is encoded by the ALG13 gene (Gene ID: 79868), which has been associated with X-linked intellectual disability, epilepsy, and congenital disorders of glycosylation (CDG) . Understanding ALG13's function is critical for researchers studying developmental disorders, as pathogenic variants in ALG13 can alter N-linked protein glycosylation in both female and male subjects . The protein has a calculated molecular weight of 18 kDa (165 amino acids), though observed weights in experimental conditions can vary considerably .

What validated applications exist for ALG13 antibodies in current research?

ALG13 antibodies have been validated for multiple laboratory applications, with evidence-based protocols available. According to product validation data, ALG13 antibodies have demonstrated effectiveness in:

Researchers should note that optimal dilutions are often sample-dependent and may require titration for best results in specific experimental systems .

How should researchers optimize ALG13 antibody protocols for different tissue types?

For brain tissue samples, which show reliable ALG13 expression, immunohistochemistry protocols require careful optimization of antigen retrieval methods. The recommended approach involves:

• Primary antigen retrieval with TE buffer at pH 9.0

• Alternative retrieval using citrate buffer at pH 6.0 if initial results are suboptimal

• Antibody dilution starting at 1:50-1:500 for IHC applications, with optimization for specific tissue types

• Proper controls including both positive (mouse brain tissue) and negative controls

For non-neural tissues, researchers should conduct preliminary validation studies as ALG13 expression patterns may vary considerably across tissue types .

What molecular variants of ALG13 should researchers consider when designing experiments?

When designing experiments to investigate ALG13, researchers should account for:

• The most frequent pathogenic variant: c.320A>G; p.(Asn107Ser), which has been reported in 37 females and two males with developmental delay and epileptic encephalopathy

• Novel variants, including three recently reported inherited variants

• Potential tissue-specific expression patterns that might affect antibody detection sensitivity

Researchers investigating ALG13-related disorders should consider that variants may produce subtle alterations in N-linked protein glycosylation that require sensitive detection methods beyond standard techniques .

What are the technical challenges in detecting ALG13 expression using antibodies?

A significant technical challenge when working with ALG13 antibodies is the discrepancy between calculated and observed molecular weights. While the calculated molecular weight is 18 kDa, observed molecular weights in experimental conditions have been reported at both 18 kDa and 130-135 kDa . This substantial difference requires researchers to:

• Run appropriate molecular weight markers covering a wide range

• Include positive controls (such as Jurkat cell lysates) to confirm band identity

• Consider post-translational modifications or protein complexes that might explain the higher molecular weight observations

• Use additional confirmation methods such as knockout/knockdown validation or mass spectrometry

How can researchers apply ALG13 antibodies to investigate glycosylation disorders?

For investigating congenital disorders of glycosylation (CDG) related to ALG13 dysfunction, researchers should consider these methodological approaches:

• Combine ALG13 antibody studies with glycan analysis techniques such as mass spectrometry

• Correlate antibody detection of ALG13 variants with transferrin glycosylation profiles

• Design experiments that can detect subtle alterations in N-glycan profiles, as ALG13 variants may cause only minor changes (e.g., 0.3-0.5% changes in total plasma glycans)

• Implement controls for gender-specific differences, as glycosylation patterns may differ between males and females with identical variants

Research has shown that among ALG13-CDG affected individuals, abnormal serum transferrin glycosylation may not be universally present, with only one male among eleven unrelated individuals showing such abnormalities .

What are the most common sources of experimental error when using ALG13 antibodies?

When using ALG13 antibodies, researchers frequently encounter these challenges:

• Non-specific binding, particularly in complex tissue samples

• Inconsistent results due to improper storage conditions (antibody should be stored at -20°C in PBS with 0.02% sodium azide and 50% glycerol at pH 7.3)

• Suboptimal signal-to-noise ratio in immunohistochemistry applications

• Difficulty in detecting minor variants or post-translational modifications

To minimize these issues, researchers should:

• Perform thorough blocking steps using appropriate blocking buffers

• Validate antibody specificity through multiple approaches (Western blot, IHC, IF)

• Consider sample preparation techniques that preserve protein conformation

• Optimize antibody concentration through careful titration experiments

How should researchers design validation experiments for ALG13 antibodies?

A robust validation strategy for ALG13 antibodies should include:

• Multi-technique validation across WB, IHC, and IF applications using the same sample sources

• Comparison of results from different antibody clones or sources when available

• Use of genetic models (knockdown or knockout) to confirm specificity

• Correlation of protein detection with mRNA expression data

• Peptide competition assays to confirm epitope specificity

For developmental disorder research, validation should include testing on samples with known ALG13 variants to establish detection sensitivity for altered forms of the protein .

How can ALG13 antibodies contribute to understanding X-linked intellectual disability?

ALG13 antibodies provide valuable tools for investigating the relationship between ALG13 variants and X-linked intellectual disability by:

• Enabling visualization of ALG13 expression patterns in neural tissues

• Facilitating comparative studies between normal and pathogenic variant expressions

• Supporting investigations into potential therapeutic targets within the N-glycosylation pathway

• Allowing correlation between protein expression and clinical phenotypes

Researchers have identified ALG13 variants using exome sequencing approaches, including through the Deciphering Developmental Disorders (DDD) Study. These genetic findings can be complemented with protein expression studies using validated ALG13 antibodies to understand functional consequences of mutations .

What considerations are important when studying ALG13 in epilepsy models?

When investigating ALG13's role in epilepsy, particularly infantile spasms/West syndrome and epileptic encephalopathy, researchers should consider:

• The specific cellular localization of ALG13 in brain tissue using optimized immunofluorescence protocols

• Potential differences in protein expression between normal and epileptic brain tissue

• Correlation between ALG13 variants and glycosylation abnormalities in epilepsy models

• Age-dependent expression patterns, particularly in developmental models

The c.320A>G; p.(Asn107Ser) variant has been significantly associated with infantile spasms and epileptic encephalopathy, making this a critical target for antibody-based research applications .

What emerging techniques might enhance ALG13 antibody applications in glycobiology research?

Emerging technologies that may enhance ALG13 antibody applications include:

• Single-cell proteomics to detect cell-specific ALG13 expression patterns

• Proximity ligation assays (PLA) to study ALG13 protein interactions in situ

• CRISPR-based genetic models combined with antibody detection for functional studies

• Advanced glycomics approaches to correlate ALG13 expression with specific glycan alterations

• Super-resolution microscopy for precise subcellular localization studies

These approaches can help address the current limitations in understanding how ALG13 variants affect N-linked glycosylation pathways in developmental disorders .

How might researchers design experiments to resolve contradictory findings about ALG13 function?

To address contradictory findings regarding ALG13 function, researchers should design experiments that:

• Compare multiple antibodies targeting different epitopes of ALG13

• Combine protein detection with functional glycosylation assays

• Utilize both in vitro and in vivo models to validate findings

• Account for tissue-specific and developmental-stage-specific variations

• Implement quantitative approaches to measure subtle changes in glycosylation patterns

Current evidence suggests that ALG13 pathogenic variants may produce only subtle alterations in N-linked protein glycosylation, which could explain contradictory findings if detection methods lack sufficient sensitivity .