slc30a6 Antibody

What is SLC30A6 and why is it significant for research?

SLC30A6, also known as ZnT6, belongs to the solute carrier family 30 (zinc transporter) member 6. This protein functions primarily as a zinc-efflux transporter that allocates cytoplasmic zinc to the trans-Golgi network (TGN) and vesicular compartments. Unlike some transporters, SLC30A6 likely doesn't possess intrinsic transporter activity independently but forms a functional zinc ion:proton antiporter heterodimer with SLC30A5 (ZnT5) .

This zinc transporter is particularly significant in research because it contributes to zinc homeostasis within the early secretory pathway and regulates the activation and folding of enzymes such as alkaline phosphatases and enzymes involved in phosphatidylinositol glycan anchor biosynthesis . Importantly, altered expression of SLC30A6 has been observed in Alzheimer's disease brain plaques, making it a valuable target for neurodegenerative disease research .

What types of SLC30A6 antibodies are available for research applications?

Several types of SLC30A6 antibodies are commercially available for various research applications:

• Region-specific antibodies:

  • N-terminal targeting antibodies (e.g., against aa 1-216)

  • Central region targeting antibodies (e.g., against aa 250-400, aa 282-310)

  • C-terminal targeting antibodies

• Host-specific production:

  • Most SLC30A6 antibodies are rabbit polyclonal antibodies

  • Different clones target various epitopes for different applications

• Application-validated antibodies:

  • Western blotting (WB) validated antibodies

  • Immunohistochemistry (IHC) validated antibodies

  • Immunofluorescence (ICC/IF) validated antibodies

  • ELISA validated antibodies

The selection of an appropriate antibody depends on the experimental goals, target species, and specific region of interest within the SLC30A6 protein structure.

What applications are SLC30A6 antibodies validated for?

SLC30A6 antibodies have been validated for multiple experimental applications according to manufacturer data:

The observed molecular weight of SLC30A6 in Western blot applications is approximately 51 kDa, which aligns with its calculated molecular weight of 50-51 kDa . When designing experiments, it's important to note that SLC30A6 localizes primarily to the Golgi apparatus and vesicular compartments, which should inform appropriate controls and co-localization studies .

Which species reactivity is available for SLC30A6 antibodies?

SLC30A6 antibodies with validated reactivity against multiple species are commercially available:

• Human: Most widely available, with numerous antibodies validated across various applications for human SLC30A6 detection .

• Mouse: Several antibodies demonstrate confirmed reactivity to mouse SLC30A6, making them suitable for murine model research .

• Rat: Some antibodies show cross-reactivity with rat SLC30A6 .

• Other species: Limited antibodies may have predicted reactivity with additional species such as:

  • Bovine

  • Chicken

  • Xenopus

When selecting antibodies for cross-species applications, sequence homology is an important consideration. Some manufacturers note high sequence identity (89%) between human SLC30A6 immunogens and mouse/rat orthologs, suggesting effective cross-reactivity . Always verify the validated species reactivity before designing experiments across different model systems.

How should I determine the optimal working dilution for SLC30A6 antibodies in Western blot applications?

Determining the optimal working dilution for SLC30A6 antibodies requires a systematic titration approach:

• Start with the manufacturer's recommended range:

  • Most SLC30A6 antibodies suggest initial dilutions of 1:500-1:2000 for Western blot applications

  • This range provides a starting point for optimization

• Perform a titration experiment:

  • Prepare identical Western blot membranes with your samples

  • Test at least 3-4 dilutions (e.g., 1:500, 1:1000, 1:2000)

  • Include positive controls (e.g., A375 cells, K-562 cells)

  • Include negative controls (e.g., SLC30A6 knockdown cells if available)

• Evaluate based on these criteria:

  • Signal-to-noise ratio: Select the dilution providing strongest specific signal with minimal background

  • Band specificity: Confirm a single band at the expected molecular weight (approximately 51 kDa for SLC30A6)

  • Reproducibility: Ensure consistent results across replicate experiments

• Consider additional optimization parameters:

  • Blocking conditions: 5% non-fat milk or BSA in TBS/PBS

  • Primary antibody incubation: Overnight at 4°C versus 2 hours at room temperature

  • Wash stringency: Number and duration of washes with 0.1% TBST

• Validation strategies:

  • Use blocking peptides when available for validation

  • Some manufacturers offer corresponding recombinant protein fragments (e.g., Human SLC30A6 (aa 290-374) Control Fragment) that can be used at 100x molar excess to confirm specificity

This methodical approach ensures optimal antibody performance and reliable detection of SLC30A6 in your experimental system.

What tissue preparation protocols are recommended for SLC30A6 immunohistochemistry studies?

Effective tissue preparation for SLC30A6 immunohistochemistry requires specific attention to fixation, antigen retrieval, and blocking:

• Fixation options:

  • Formalin fixation followed by paraffin embedding (FFPE):

    • Fix tissues in 10% neutral buffered formalin for 24-48 hours

    • Process and embed in paraffin following standard histology protocols

  • Fresh-frozen sections:

    • Flash freeze tissue in optimal cutting temperature (OCT) compound

    • Store at -80°C until sectioning

• Sectioning parameters:

  • FFPE tissues: 4-6 μm thick sections mounted on positively charged slides

  • Frozen sections: 8-10 μm thick sections on coated slides

• Antigen retrieval (critical for FFPE sections):

  • Heat-induced epitope retrieval (HIER):

    • Citrate buffer (pH 6.0): Heat to 95-100°C for 15-20 minutes

    • EDTA buffer (pH 9.0): Alternative if citrate buffer yields suboptimal results

  • Allow slides to cool gradually to room temperature (20-30 minutes)

• Permeabilization:

  • For membrane proteins like SLC30A6, include a permeabilization step

  • Use 0.1-0.3% Triton X-100 in PBS for 10-15 minutes at room temperature

• Blocking and antibody incubation:

  • Block with 5-10% normal serum (from secondary antibody host species)

  • Include 1% BSA to reduce nonspecific binding

  • Dilute primary SLC30A6 antibody appropriately (typically 1:100-1:500)

  • Incubate overnight at 4°C in a humidified chamber

  • For secondary antibody detection, use HRP or fluorophore-conjugated antibodies

For brain tissue specifically (where SLC30A6 is highly expressed), additional considerations include extended fixation times for proper tissue penetration and more aggressive antigen retrieval. Always include positive control tissues with known SLC30A6 expression (e.g., cerebellum, hippocampus) to validate staining protocols .

How can I validate the specificity of SLC30A6 antibodies in my experimental system?

Validating antibody specificity is critical for ensuring reliable SLC30A6 detection. A comprehensive validation approach includes:

• Positive and negative controls:

  • Positive tissue/cell controls: Use samples with confirmed SLC30A6 expression (e.g., brain tissues, A375, or K-562 cells)

  • Negative controls: Use SLC30A6 knockout or knockdown samples (siRNA/shRNA treated)

  • Isotype controls: Use matched isotype antibodies to assess non-specific binding

• Peptide competition assays:

  • Pre-incubate the antibody with excess immunizing peptide

  • Run parallel experiments with blocked and unblocked antibody

  • Signal elimination in the peptide-blocked sample confirms specificity

  • Manufacturers recommend a 100x molar excess of the protein fragment control

• Multiple antibody validation:

  • Test antibodies targeting different epitopes of SLC30A6 (N-terminal vs. C-terminal)

  • Concordant results from antibodies targeting different regions increase confidence

  • Compare results using antibodies from different vendors that target the same protein

• Western blot validation:

  • Confirm single band at expected molecular weight (51 kDa)

  • Verify band disappearance in knockdown/knockout samples

  • Compare with published literature for band pattern consistency

• Subcellular localization confirmation:

  • SLC30A6 should localize to Golgi apparatus and vesicular compartments

  • Co-staining with organelle markers (e.g., Golgi markers) should show appropriate colocalization

  • Aberrant localization patterns may indicate antibody cross-reactivity

This systematic approach ensures that observed signals genuinely represent SLC30A6 and not cross-reactive or non-specific binding, which is particularly important when studying zinc transporter family members with structural similarities.