This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
HAL / Histidine ammonia-lyase · Western blot design guide
Design a Western Blot for HAL
Real validated HAL Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-HAL WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Evidence assembled September 2026 · For research use; verify linked source records and product datasheet before use
HAL Western Blot Experimental Design Guide
Expected bands, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~72.7 kDa | |
| Observed band | ~75 kDa | |
| Gel | 10% (catalog A02612-1) | |
| Positive control | Liver (IHC candidate; verify WB) +4 more | |
| Negative control | Ovary (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 3 isoform(s) |
Section 1
Real Curated HAL Western Blot Protocols
The A02612-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | rat liver, mouse liver (catalog A02612-1) |
| Gel % | 10% (catalog A02612-1) |
| Load | 30 ug; reducing conditions (catalog A02612-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A02612-1) |
| Membrane | nitrocellulose membrane (catalog A02612-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A02612-1) |
| Primary antibody | A02612-1 · 0.5 μg/mL (catalog A02612-1) |
| Primary incubation | overnight at 4°C (catalog A02612-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A02612-1) |
| Secondary incubation | 1.5 hour at RT (catalog A02612-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A02612-1) |
| Detection | ECL (catalog A02612-1) |
Section 2
What Is the Expected HAL Western Blot Band Size?
HAL is predicted at 72.7 kDa and observed at ~75 kDa in reducing liver lysates; the cause of the difference is not established.
| Band at ~75 kDa | Empirical HAL band in reducing liver lysates; confirm identity with controls. |
| Band near 72.7 kDa | Consistent with the sequence-predicted mass of HAL. |
| Doublet near the main band | Could reflect HAL forms, but the supplied evidence does not establish their migration. |
| Several bands at different masses | Isoforms 1, 2, and 3 exist, but distinct resolved bands have not been demonstrated. |
💡Expected HAL appearanceHAL has a predicted mass of 72.7 kDa and an empirical band at ~75 kDa in reducing liver lysates; confirm band identity with appropriate controls.
| UniProt predicted mass | 72.7 kDa is the sequence-based reference; the observed band is ~75 kDa. |
| Splice isoform 1 | Its individual mass and migration are not supplied. |
| Splice isoform 2 | Its size relative to isoform 1 and its migration are not supplied. |
| Splice isoform 3 | Its size relative to the other isoforms and its migration are not supplied. |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | The ~75 kDa empirical band runs above the 72.7 kDa prediction; the cause is unestablished. | Compare with the documented liver lysate band and use an independent HAL antibody or depletion control. |
| Band lower than expected | An isoform or fragment is possible, but its identity is unestablished. | Check with an independent HAL antibody and compare sample preparation conditions. |
| Multiple bands | HAL has three isoforms, but their band positions are unknown. | Test band identity with an independent antibody or HAL depletion control. |
| Weak or no signal | HAL abundance or antibody detection may be insufficient in the sample. | Include a liver lysate positive control and verify loading and transfer. |
| Fragments below expected size | Sample degradation is possible; no fragment masses are supplied. | Prepare fresh lysate with protease inhibitors and check band identity with an independent antibody. |
Experiment evidence
Western blot images showing HAL band
A02612-1 · HAL
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for HAL Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for HAL in Western blot, you can use liver tissue, where HPA reports high expression.
Positive control: Liver (IHC candidate; verify WB)
Negative control: Ovary (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: High expression in liver and no detected expression in ovary make tissue controls feasible.
HPA tissue expression evidence for HAL
Comprehensive Human Protein Atlas IHC scoring per tissue. Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Higher expression tissues · candidate positive controls from IHC
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Liver | hepatocytes | High | Protein (IHC) | HPA → |
| Skin | cells in basal layer | High | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | glandular cells | Medium | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Ovary | ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Caudate | neuronal cells | Low | Protein (IHC) | HPA → |
| Hippocampus | neuronal cells | Low | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced HAL Western Blot Tips
Deeper troubleshooting and optimisation questions for HAL, answered from its protein features.
How should HAL band migration be interpreted?
Band shift · Use the separately labelled calculated mass and catalog-observed evidence above. A sequence annotation does not establish an observed migration shift. Verify target identity with orthogonal controls.
Could HAL isoforms produce different bands?
Isoforms · Yes. UniProt lists three isoforms. Isoform 3 lacks canonical residues 1–208; isoform 2 replaces canonical residues 589–657 with FGK. Both sequence changes could affect band position. Confirm any additional band with an isoform-specific reagent or sequence-based assay.
For detection across isoforms, choose an epitope outside canonical residues 1–208 and 589–657. An epitope within 1–208 would miss isoform 3; one within 589–657 may miss isoform 2. Check the antibody’s stated epitope before comparing bands.
For detection across isoforms, choose an epitope outside canonical residues 1–208 and 589–657. An epitope within 1–208 would miss isoform 3; one within 589–657 may miss isoform 2. Check the antibody’s stated epitope before comparing bands.
Which HAL modifications matter when interpreting bands?
PTM · UniProt lists 2,3-didehydroalanine from Ser254 and phosphorylation at Thr396, Ser635, Thr637, and Ser648. These are canonical UniProt coordinates. The tail replacement in isoform 2 removes the canonical sequence containing the last three sites. Modification alone does not establish a visible shift.
It can test whether phosphorylation contributes to a band pattern, given the listed phosphosites at Thr396, Ser635, Thr637, and Ser648. Compare matched treated and untreated samples. A change would support a phosphorylation contribution, but would not by itself assign the responsible site.
It can test whether phosphorylation contributes to a band pattern, given the listed phosphosites at Thr396, Ser635, Thr637, and Ser648. Compare matched treated and untreated samples. A change would support a phosphorylation contribution, but would not by itself assign the responsible site.
Does this guide establish induction of HAL?
Induction · No general induction response is established by this guide. A pathway or gene-set association is not evidence of induction in a particular specimen. Verify the relevant treatment and control in a target-specific experiment.
How should transfer be checked for HAL?
Transfer · Standard workflow guidance: verify transfer efficiency for the intended target size before interpreting a weak signal. Use total-protein assessment and optimize transfer for the membrane, gel and apparatus; the labelled catalog values take precedence.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A02612-1 datasheet where specified. Otherwise compare 5% milk or 5% BSA in TBST; for a phospho-specific assay start with BSA. Optimize background and specific signal with matched controls.
How should HAL bands be quantified across samples?
Quantitation · Quantify the same verified HAL band across samples and record whether the antibody detects all three isoforms. Isoform 3 lacks residues 1–208, and isoform 2 has a replacement at 589–657; an antibody targeting either region may give unequal isoform coverage.
Why does HAL appear near 75 kDa instead of 72.7 kDa?
Interpretation · The reported band is approximately 75 kDa, while the predicted mass is 72.7 kDa. The listed features do not establish the cause of this difference. Use the 75 kDa band as an empirical reference and verify its identity with a HAL-specific control.
Check whether their positions and antibody recognition are consistent with isoforms 2 or 3, then test HAL identity independently. Consider the listed phosphosites when evaluating nearby bands, but do not assign a band to a specific modification from mobility alone. UniProt lists no glycosylation sites for HAL.
Check whether their positions and antibody recognition are consistent with isoforms 2 or 3, then test HAL identity independently. Consider the listed phosphosites when evaluating nearby bands, but do not assign a band to a specific modification from mobility alone. UniProt lists no glycosylation sites for HAL.
Boster reagents
HAL Western Blot Antibodies
Catalog antibodies with Western blot application and product-specific WB images. Evaluate suitability with the reported sample, controls and experimental conditions.
Real WB data
Anti-HAL Antibody Picoband®
Cat # A02612-1
The catalog reports one anti-HAL antibody, A02612-1, with stated Human, Mouse, and Rat reactivity. Its Western blot image uses rat and mouse liver lysates and shows a band near 75 kDa, close to the expected 73 kDa. No human sample is shown.
Which to pick: A02612-1 is the only listed choice. Its WB image provides an example for rat and mouse liver lysates at 0.5 μg/mL antibody. Human reactivity is listed, but the supplied image does not test a human sample.
Source: BosterBio HAL gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P42357.
- Human Protein Atlas. HAL tissue expression.
- PMC12662978 — target-verified WB comparison