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- Table of Contents
KMT2B / Histone-lysine N-methyltransferase 2B · Western blot design guide
Design a Western Blot for KMT2B
Source-linked KMT2B Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-KMT2B 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
KMT2B 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 | ~293.5 kDa | |
| Gel | 4–12% gradient (standard starting point) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked KMT2B Western Blot Protocol Options
The A05680 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | A549, NIH-3T3, H9C2 (catalog A05680) |
| Gel % | 4–12% gradient (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Wet/tank transfer; optimize duration (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A05680; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Species-matched HRP conjugate at validated dilution (standard starting point) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL; bracket exposures to avoid saturation (standard starting point) |
Section 2
What Is the Expected KMT2B Western Blot Band Size?
KMT2B is predicted at 293.5 kDa; listed modifications may affect migration, but no empirical band or visible shift is established.
| Band near 293.5 kDa | Consistent with predicted full-length KMT2B; confirm band identity |
| Weak band in whole-cell lysate | Nuclear KMT2B may be diluted by other cellular proteins |
| Stronger band in nuclear extract | Consistent with KMT2B nuclear localization |
| Nearby doublet | Listed phosphorylation sites could contribute, but a visible shift is unproven |
💡Expected KMT2B appearanceKMT2B has a predicted mass of 293.5 kDa, but no empirical band size is supplied; confirm a candidate band with identity controls.
| Predicted molecular mass | Places full-length KMT2B near 293.5 kDa; migration is unverified |
| N-acetylalanine at residue 2 | Adds a small modification; a visible size shift is unproven |
| Phosphoserines at residues 113, 114, and 118 | May affect migration; a resolved shift is unproven |
| Phosphoserines at residues 821, 844, and 861 | May affect migration; a resolved shift is unproven |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | Nuclear KMT2B may be underrepresented in the preparation | Check nuclear extraction and a positive control |
| Band higher than expected | Band identity or apparent migration is uncertain | Check molecular-weight markers and an independent antibody |
| Band lower than expected | Band identity or protein integrity is uncertain | Check an independent antibody and sample integrity |
| Multiple bands | Specificity is uncertain; listed phosphorylation sites do not establish a resolved pattern | Compare an independent antibody and test phosphatase sensitivity |
| Weak or no signal | KMT2B may be underrepresented in whole-cell lysate | Enrich the nuclear fraction and check a positive control |
| Fragments below expected size | Sample degradation is possible | Use protease inhibitors and compare fresh lysate |
Experiment evidence
Western blot images showing KMT2B band
A05680 · KMT2B
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for KMT2B Western blot
🧪For positive controls for KMT2B in Western blot, you can use no HPA-validated tissue or cell line from the supplied data.
Positive control: No high/medium HPA tissue identified
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: HPA data are unavailable, so tissue controls cannot be identified; use a knockdown or KO line to validate the nuclear signal.
HPA tissue expression evidence for KMT2B
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 |
|---|---|---|---|---|
| No high/medium HPA tissues identified in the supplied evidence. | ||||
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| No lower-expression tissue rows available in the supplied evidence. | ||||
Section 3
Advanced KMT2B Western Blot Tips
Deeper troubleshooting and optimisation questions for KMT2B, answered from its protein features.
How should KMT2B band size compare with its predicted mass?
Band shift · The supplied sequence predicts 293.5 kDa. No empirical apparent band is available, so use 293.5 kDa as a reference, not an expected exact migration position. The listed modifications alone do not establish a visible shift.
Could a second KMT2B band represent an isoform?
Isoforms · The supplied record lists one isoform and no alternative sequence. It provides no basis to assign a second band to a KMT2B isoform; check the band’s identity before interpreting it.
Which KMT2B modifications matter when interpreting band shifts?
PTM · UniProt lists N-acetylalanine at position 2; phosphoserine at 113, 114, 118, 351, 821, 844, 861, 936, 1032, 1035, 1092, 1095, 1930, 1936, 2288 and 2348; and phosphothreonine at 2068 and 2083. These are UniProt coordinates and may differ from antibody or paper numbering. Their presence does not demonstrate a visible shift.
Does this guide establish induction of KMT2B?
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.
What transfer method to use for KMT2B Western blot?
Transfer · At a predicted 293.5 kDa, KMT2B is large. Start with a wet transfer configured for high molecular weight proteins and verify that the region around 293.5 kDa transfers effectively before interpreting a weak or absent band.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A05680 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 KMT2B signal be quantified?
Quantitation · KMT2B is listed as nuclear. Compare samples prepared in the same way and normalize against a suitable nuclear loading measure. Quantify the same validated KMT2B band across samples; the supplied features do not establish that bands at different positions represent equivalent amounts of protein.
How should unexpected KMT2B bands be interpreted?
Interpretation · Check whether the band tracks with nuclear KMT2B and verify its identity with an independent antibody or target depletion. The record lists one isoform, no signal peptide or propeptide, and no empirical band position. Phosphorylation is documented, but the listed sites alone cannot explain a particular extra band.
Boster reagents
KMT2B 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-WBP7 KMT2B Antibody
Cat # A05680
A05680 is a KMT2B antibody listed for human, mouse, and rat. Its WB image shows A549, NIH-3T3, and H9C2 whole-cell lysates at 1:500 dilution. No publication evidence was supplied.
Which to pick: A05680 is the only listed option. It has a WB image for the three named cell lysates; choose it based on your sample and the reported 1:500 condition.
Source: BosterBio KMT2B gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9UMN6.
- Human Protein Atlas. KMT2B tissue expression.
- PMC10863086 — target-verified WB comparison
- PMC10493599 — target-verified WB comparison
- PMC12783180 — target-verified WB comparison