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- Table of Contents
MIS12 / Protein MIS12 homolog · Western blot design guide
Design a Western Blot for MIS12
Source-linked MIS12 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-MIS12 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
MIS12 Western Blot Experimental Design Guide
Expected bands, source-linked protocol options, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~24.1 kDa | |
| Observed band | ~25 kDa | |
| Gel | 5–20% (catalog A06832-1) | |
| Positive control | Lymph node (IHC candidate; verify WB) +4 more | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | — | |
| Caveat | — | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked MIS12 Western Blot Protocol Options
The A06832-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human Hela, human 293T, human HepG2, human Jurkat, human Hacat, human U251, human SH-SY5Y, human K562 (catalog A06832-1) |
| Gel % | 5–20% (catalog A06832-1) |
| Load | 30 ug; reducing conditions (catalog A06832-1) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A06832-1) |
| Membrane | nitrocellulose membrane (catalog A06832-1) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A06832-1) |
| Primary antibody | A06832-1 · 0.25 μg/mL (catalog A06832-1) |
| Primary incubation | overnight at 4°C (catalog A06832-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A06832-1) |
| Secondary incubation | 1.5 hour at RT (catalog A06832-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A06832-1) |
| Detection | ECL (catalog A06832-1) |
Section 2
What Is the Expected MIS12 Western Blot Band Size?
MIS12 is predicted at 24.1 kDa and observed near 25 kDa; the cause of the difference is not established by the supplied features.
| Single band near 25 kDa | Matches the reported MIS12 band in whole-cell lysates. |
| Band near 24.1 kDa | Consistent with the predicted mass of intact MIS12. |
| No band near 25 kDa | MIS12 was not detected; check recovery and assay sensitivity. |
| Additional bands away from 25 kDa | Their identity is not established by the supplied MIS12 features. |
💡Expected MIS12 appearanceMIS12 has a predicted mass of 24.1 kDa and a reported band near 25 kDa in reducing whole-cell blots; confirm identity with an appropriate positive control or MIS12 depletion.
| Predicted molecular mass | Places intact MIS12 near 24.1 kDa. |
| 205-amino-acid sequence | Underlies the predicted 24.1 kDa mass; no separate size shift is established. |
| Single annotated isoform | Provides no evidence for isoform-dependent band sizes. |
| MIS12 complex membership | Does not establish a different apparent mass for MIS12 on the reported reducing blot. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | MIS12 may be poorly recovered from its kinetochore-associated compartment or below detection. | Check sample preparation and a lysate known to yield the near-25 kDa band. |
| Band higher than expected | No annotated MIS12 modification explains the higher position. | Compare with the reported near-25 kDa band and verify identity by MIS12 depletion. |
| Band lower than expected | No annotated cleavage explains the lower position. | Check sample integrity and whether the band decreases after MIS12 depletion. |
| Multiple bands | Only one isoform is annotated; additional band identities are unestablished. | Compare bands across a positive control and MIS12-depleted sample. |
| Weak or no signal | MIS12 detection may be limited by sample recovery or assay sensitivity. | Check loading, transfer and antibody performance with a positive lysate. |
Experiment evidence
Western blot images showing MIS12 band
A06832-1 · MIS12
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for MIS12 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for MIS12 in Western blot, you can use lymph node lysate, which HPA rates as highly positive.
Positive control: Lymph node (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain such as Ponceau alongside the samples.
⚠️Feasibility: Adipose tissue is an HPA not-detected comparator, but confirm its signal in your lysate.
HPA tissue expression evidence for MIS12
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 |
|---|---|---|---|---|
| Lymph node | non-germinal center cells | High | Protein (IHC) | HPA → |
| Tonsil | non-germinal center cells | High | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Caudate | neuronal cells | Medium | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | adipocytes | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Hippocampus | glial cells | Not detected | Protein (IHC) | HPA → |
| Lung | alveolar cells | Not detected | Protein (IHC) | HPA → |
| Oral mucosa | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced MIS12 Western Blot Tips
Deeper troubleshooting and optimisation questions for MIS12, answered from its protein features.
How should MIS12 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.
Should MIS12 isoforms produce multiple bands?
Isoforms · Only one isoform is listed, with no alternative sequence supplied. These features do not support assigning additional bands to MIS12 isoforms.
Could annotated modifications explain a MIS12 band shift?
PTM · No modified residues or glycosylation sites are listed. The supplied features therefore do not support attributing a shifted band to a specific modification.
Does this guide establish induction of MIS12?
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 MIS12?
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 A06832-1 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 MIS12 bands be quantified across samples?
Quantitation · Quantify the same approximately 25 kDa band across samples prepared in the same way. Because MIS12 is kinetochore-associated and involved in cell division, account for differences in sample composition and cell-cycle state when interpreting changes.
Why is MIS12 observed near 25 kDa instead of 24.1 kDa?
Interpretation · The observed band is approximately 25 kDa, close to the predicted 24.1 kDa for the 205-residue protein. The supplied features do not establish a specific cause for the small difference in apparent mass.
MIS12 is associated with the chromosome centromere and kinetochore. Keep the sample fraction consistent across lanes so differences in MIS12 recovery do not complicate comparison.
MIS12 belongs to a complex with DSN1, NSL1 and PMF1 and interacts with several other proteins. These associations alone do not identify an unexpected Western-blot band as MIS12. Compare it with the approximately 25 kDa band before assigning its identity.
MIS12 is associated with the chromosome centromere and kinetochore. Keep the sample fraction consistent across lanes so differences in MIS12 recovery do not complicate comparison.
MIS12 belongs to a complex with DSN1, NSL1 and PMF1 and interacts with several other proteins. These associations alone do not identify an unexpected Western-blot band as MIS12. Compare it with the approximately 25 kDa band before assigning its identity.
Boster reagents
MIS12 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-MIS12 Antibody Picoband®
Cat # A06832-1
The catalog reports one anti-MIS12 antibody for Western blotting, A06832-1, with reported human reactivity. Its WB image shows a band near 25 kDa, close to the expected 24 kDa, across eight human cell lysates. Evidence is limited to the supplied product image and caption.
Which to pick: A06832-1 is the only listed option. Its WB caption documents human cell lysates and an antibody concentration of 0.25 μg/mL, making those conditions the practical starting point for a similar sample.
Source: BosterBio MIS12 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9H081.
- Human Protein Atlas. MIS12 tissue expression.
- PMC10961024 — target-verified WB comparison
- PMC6586897 — target-verified WB comparison
- PMC6930117 — target-verified WB comparison