SLC19A2 Antibody, FITC conjugated

What is SLC19A2 and why is it significant in research?

SLC19A2 encodes thiamine transporter 1 (THTR1), a high-affinity transporter for the intake of thiamine across the cell membrane. This protein has gained significant research interest because mutations in the SLC19A2 gene cause thiamine-responsive megaloblastic anemia syndrome (TRMA), an autosomal recessive disorder characterized by diabetes mellitus, megaloblastic anemia, and sensorineural deafness . Recent research has also linked SLC19A2 deficiency to impaired insulin secretion in conjunction with mitochondrial dysfunction, loss of protection against oxidative stress, and cell cycle arrest, making it a critical target for diabetes research .

What are the key characteristics of SLC19A2 Antibody, FITC conjugated?

The SLC19A2 Antibody, FITC conjugated is a rabbit polyclonal antibody directed against SLC19A2, conjugated to fluorescein isothiocyanate (FITC). The antibody is generated using a recombinant Human Thiamine transporter 1 protein fragment (amino acids 209-285) as the immunogen . It specifically reacts with human samples and has been validated for applications including ELISA and Dot Blot techniques . The antibody is supplied in liquid form, typically in a preservative buffer containing 50% glycerol, 0.01M PBS, pH 7.4, and 0.03% Proclin 300 to maintain stability during storage .

What protein aliases and gene synonyms are associated with SLC19A2?

SLC19A2 is known by several aliases in the scientific literature and databases. Protein aliases include high affinity thiamine transporter, reduced folate carrier protein (RFC) like, solute carrier family 19 (thiamine transporter) member 2, Solute carrier family 19 member 2, TC1, Thiamine carrier 1, Thiamine transporter 1, and ThTr-1 . Gene aliases include SLC19A2, TC1, THMD1, THT1, THTR1, and TRMA . The protein has the UniProt ID O60779 and Entrez Gene ID 10560 in humans . Understanding these alternative nomenclatures is essential when conducting literature searches or database queries related to this transporter.

What are the validated applications for SLC19A2 Antibody, FITC conjugated?

The SLC19A2 Antibody, FITC conjugated has been validated primarily for ELISA and Dot Blot applications . When considering related non-FITC conjugated SLC19A2 antibodies, the range of applications expands to include Western Blot, Immunohistochemistry (IHC), Immunocytochemistry (ICC), and Immunofluorescence (IF) . For immunohistochemistry applications specifically, a recommended dilution range of 1:200-1:500 has been established . Each application requires specific optimization protocols, and researchers should validate the antibody in their specific experimental systems before proceeding with large-scale experiments.

How should SLC19A2 Antibody, FITC conjugated be stored and handled for optimal performance?

For optimal performance and longevity, SLC19A2 Antibody, FITC conjugated should be stored at -20°C or -80°C immediately upon receipt . Repeated freeze-thaw cycles should be avoided as they can compromise antibody integrity and function . When working with the antibody, it's recommended to aliquot the stock solution into smaller volumes for single use to minimize freeze-thaw cycles. Since the antibody is FITC-conjugated, it's light-sensitive, so researchers should protect it from prolonged exposure to light during handling and storage to prevent photobleaching of the fluorophore.

What experimental controls should be included when using SLC19A2 Antibody, FITC conjugated?

For rigorous experimental design, several controls should be included when using SLC19A2 Antibody, FITC conjugated:

• Positive control: Cells or tissues known to express SLC19A2 (such as human pancreatic β-cells which show functional consequences of SLC19A2 deficiency)

• Negative control: Cells or tissues known not to express SLC19A2 or SLC19A2-knockdown/knockout samples

• Isotype control: A FITC-conjugated non-specific rabbit IgG at the same concentration as the SLC19A2 antibody to assess non-specific binding

• Secondary antibody-only control (for indirect detection methods): To evaluate background fluorescence

• Blocking peptide control: Pre-incubation of the antibody with the immunogen peptide (amino acids 209-285 of SLC19A2) to confirm specificity

What factors might affect the sensitivity and specificity of SLC19A2 Antibody, FITC conjugated?

Several factors can influence the performance of SLC19A2 Antibody, FITC conjugated:

• Sample preparation: Proper fixation and permeabilization are critical for antibody access to the target epitope

• Antibody dilution: Suboptimal dilutions may result in weak signals or high background

• Incubation conditions: Temperature, time, and buffer composition can affect binding kinetics

• Cross-reactivity: While the antibody is reported to be specific for human SLC19A2, potential cross-reactivity with related proteins should be considered

• Epitope accessibility: The antibody targets amino acids 209-285, which may be inaccessible in certain experimental conditions or sample preparations

• FITC photobleaching: Extended exposure to light can reduce signal intensity

Researchers should optimize these parameters for their specific experimental systems to achieve optimal results.

How can researchers troubleshoot weak or absent fluorescence signals when using SLC19A2 Antibody, FITC conjugated?

When encountering weak or absent signals with SLC19A2 Antibody, FITC conjugated, consider the following troubleshooting approaches:

• Verify SLC19A2 expression: Confirm target expression in your samples using alternative methods (RT-PCR, western blot with a different antibody)

• Optimize antibody concentration: Test a range of dilutions to determine optimal concentration

• Extend incubation time: Longer incubation at 4°C may improve signal strength

• Improve permeabilization: Ensure adequate permeabilization for intracellular antigens

• Enhance signal detection: Use anti-FITC secondary antibodies or signal amplification systems

• Check for photobleaching: Minimize light exposure during sample processing and examination

• Assess buffer compatibility: Ensure buffers do not contain components that might interfere with antibody binding

• Evaluate microscope settings: Optimize exposure settings and filter sets for FITC detection

What are the potential cross-reactivity concerns with SLC19A2 Antibody, FITC conjugated?

• Closely related proteins: SLC19A1 (reduced folate carrier) and SLC19A3 (another thiamine transporter) share sequence similarities with SLC19A2

• Species homologs: Cross-reactivity with SLC19A2 from other species may occur, although this needs to be empirically determined

• Proteins containing similar epitopes: Proteins with sequence homology to the immunogen region (amino acids 209-285) may potentially cross-react

To assess cross-reactivity, researchers can perform competitive binding assays with recombinant SLC19A2 and related proteins, or test the antibody in systems where expression of specific targets has been genetically manipulated.

How can SLC19A2 Antibody, FITC conjugated be used to investigate TRMA pathophysiology?

SLC19A2 Antibody, FITC conjugated offers valuable approaches for investigating TRMA pathophysiology:

• Cellular localization studies: Visualize membrane localization of wild-type versus mutant SLC19A2 proteins in patient-derived cells

• Functional studies: Compare thiamine uptake in cells expressing normal versus mutant SLC19A2, correlating with protein expression detected by the antibody

• Genotype-phenotype correlations: Analyze SLC19A2 expression in samples from patients with different mutations and clinical presentations

• Co-localization studies: Combine with markers for mitochondria or oxidative stress to investigate mechanisms underlying β-cell dysfunction in TRMA

• Therapy monitoring: Assess changes in SLC19A2 expression or localization in response to thiamine supplementation

These approaches can provide insights into how specific mutations affect SLC19A2 expression, localization, and function, contributing to our understanding of TRMA pathogenesis.

What methodologies can be used to study the role of SLC19A2 in diabetes and insulin secretion?

Research has linked SLC19A2 deficiency to impaired insulin secretion, suggesting a role in diabetes pathophysiology . Investigators can employ the following methodologies:

• Pancreatic β-cell imaging: Use SLC19A2 Antibody, FITC conjugated to visualize SLC19A2 expression and localization in β-cells

• Flow cytometry: Quantify SLC19A2 expression levels in isolated β-cells from various models

• Co-immunoprecipitation: Identify protein-protein interactions involving SLC19A2 in β-cells

• FACS sorting: Isolate SLC19A2-expressing cells for transcriptomic or proteomic analysis

• Live-cell imaging: Monitor thiamine transport dynamics in β-cells using the FITC-conjugated antibody for surface labeling

• Correlation studies: Relate SLC19A2 expression levels to insulin secretion capacity in various experimental conditions

These approaches can help elucidate the mechanistic link between SLC19A2 function, thiamine transport, and insulin secretion in the context of diabetes research.

How can researchers use SLC19A2 Antibody, FITC conjugated in combination with genetic analysis?

Integration of SLC19A2 Antibody, FITC conjugated with genetic analysis provides powerful approaches for comprehensive research:

• Genotype-phenotype correlations: Analyze SLC19A2 expression and localization in cells with different SLC19A2 mutations identified through sequencing

• CRISPR/Cas9 studies: Validate the specificity of antibody binding in SLC19A2 knockout models

• Mutation verification: Confirm the functional consequences of novel SLC19A2 mutations at the protein level

• Allele-specific expression: Combine with genetic markers to assess differential expression of wild-type versus mutant alleles in heterozygous samples

• Family studies: Correlate SLC19A2 expression patterns with inheritance patterns of TRMA or diabetes in families with SLC19A2 mutations

For genetic analysis, researchers can follow established protocols for DNA extraction from blood, PCR amplification of SLC19A2 exons and intron-exon junctions, and DNA sequencing as described in the literature .

How should researchers quantify and normalize fluorescence data from SLC19A2 Antibody, FITC conjugated experiments?

For rigorous quantification of fluorescence data from SLC19A2 Antibody, FITC conjugated experiments, researchers should:

• Use appropriate imaging software: ImageJ, CellProfiler, or commercial software with fluorescence quantification capabilities

• Establish consistent acquisition parameters: Maintain identical exposure times, gain settings, and acquisition parameters across all samples

• Perform background subtraction: Correct for autofluorescence using unstained controls

• Normalize to reference markers: Co-stain with markers of specific cellular compartments or use counterstains like DAPI for nuclei

• Apply appropriate statistical methods: Use measures of central tendency and dispersion appropriate for the data distribution

• Consider multiple normalization strategies:

  • Per cell normalization: Divide total signal by cell number

  • Area-based normalization: Calculate signal intensity per unit area

  • Reference gene normalization: Normalize to housekeeping proteins

What are the potential pitfalls in interpreting SLC19A2 localization data?

When interpreting SLC19A2 localization data using FITC-conjugated antibodies, researchers should be aware of several potential pitfalls:

• Fixation artifacts: Different fixation methods can alter protein localization or epitope accessibility

• Overexpression effects: In transfection studies, protein overexpression may cause aberrant localization

• Resolution limitations: Standard fluorescence microscopy may not distinguish between closely adjacent structures

• Antibody accessibility issues: The antibody may not efficiently access all cellular compartments

• Spectral overlap: FITC signal may overlap with autofluorescence or other fluorophores

• Dynamic processes: SLC19A2 may shuttle between different cellular compartments, and fixed samples capture only a static moment

• Disease state variations: SLC19A2 localization may change in disease states or in response to thiamine levels

To address these issues, researchers should use complementary approaches, appropriate controls, and consider super-resolution microscopy techniques for detailed localization studies.

How can contradictory results between SLC19A2 protein detection and functional assays be reconciled?

When faced with discrepancies between SLC19A2 protein detection using the antibody and functional thiamine transport assays, researchers can:

• Assess antibody epitope integrity: Determine if the epitope (amino acids 209-285) remains accessible in functional proteins

• Consider post-translational modifications: These may affect antibody binding but not function, or vice versa

• Evaluate protein trafficking: SLC19A2 may be detected by the antibody but not properly localized to the cell membrane for function

• Examine assay sensitivity differences: Antibody detection might be more or less sensitive than functional assays

• Analyze mutant proteins: Some mutations may allow antibody binding but impair function, as seen with the p.Lys355Gln mutation

• Check for dominant-negative effects: In heterozygous states, mutant proteins might interfere with wild-type function while still being detected by the antibody

• Consider compensatory mechanisms: Other thiamine transporters might maintain function despite reduced SLC19A2 expression

Understanding these potential disconnects between protein detection and function is particularly important when studying disease-associated mutations or therapeutic interventions.