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- Table of Contents
MAK / Serine/threonine-protein kinase MAK · Western blot design guide
Design a Western Blot for MAK
Real validated MAK Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-MAK 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
MAK 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 | ~70.6 kDa | |
| Observed band | ~70 kDa | |
| Gel | 5–20% (catalog A00407-1) | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 3 isoform(s) |
Section 1
Real Curated MAK Western Blot Protocols
The A00407-1 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human HepG2, human MCF-7 (catalog A00407-1) |
| Gel % | 5–20% (catalog A00407-1) |
| Load | 50ug; reducing conditions (catalog A00407-1) |
| Transfer | a Nitrocellulose membrane at 150mA for 50-90 minutes (catalog A00407-1) |
| Membrane | Nitrocellulose membrane (catalog A00407-1) |
| Blocking | 5% Non-fat Milk/ TBS for 1.5 hour at RT (catalog A00407-1) |
| Primary antibody | A00407-1 · 0.5 μg/mL (catalog A00407-1) |
| Primary incubation | overnight at 4°C (catalog A00407-1) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:10000 (catalog A00407-1) |
| Secondary incubation | 1.5 hour at RT (catalog A00407-1) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A00407-1) |
| Detection | ECL (catalog A00407-1) |
Section 2
What Is the Expected MAK Western Blot Band Size?
MAK is predicted at 70.6 kDa and observed at ~70 kDa; the small difference has no established cause in the supplied evidence.
| Band at ~70 kDa | Matches the empirical MAK band in reducing whole-cell lysates. |
| Single band near 70.6 kDa | Consistent with the predicted mass of MAK. |
| Additional bands at different sizes | Could reflect MAK isoforms 1, 2, or 3; their migration has not been established. |
| Doublet near ~70 kDa | Could reflect different phosphorylation states at Thr157 or Tyr159; a visible shift has not been demonstrated. |
💡Expected MAK appearanceMAK has a predicted mass of 70.6 kDa and an empirical band at ~70 kDa in reducing whole-cell lysates; confirm band identity with appropriate antibody controls.
| Predicted MAK mass | 70.6 kDa predicts a band near the empirical ~70 kDa band. |
| Isoforms 1, 2, and 3 | May differ in size, but distinct band positions are not established. |
| Autophosphorylation at Thr157 | May affect migration; a visible size shift is not established. |
| Autophosphorylation at Tyr159 | May affect migration; a visible size shift is not established. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | MAK localizes to nuclear and cytoskeletal structures that may extract poorly. | Check nuclear and cytoskeletal fractions alongside whole-cell lysate. |
| Band higher than expected | Phosphorylation could alter migration, but the cause of a higher band is unproven. | Compare phosphatase-treated samples and confirm band identity. |
| Band lower than expected | An isoform or degradation could account for a smaller band; its identity is unproven. | Check sample integrity and confirm the band with an independent MAK antibody. |
| Multiple bands | MAK has three named isoforms and two autophosphorylation sites; distinct migration is unproven. | Compare phosphatase-treated samples and verify bands with an independent MAK antibody. |
| Weak or no signal | MAK may be poorly recovered from its nuclear or cytoskeletal locations. | Check extraction and loading with fraction markers and a positive control. |
Experiment evidence
Western blot images showing MAK band
A00407-1 · MAK
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for MAK Western blot
🧪For positive controls for MAK in Western blot, you can use no HPA-supported tissue or cell line because no expression data were supplied.
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 MAK.
⚠️Feasibility: HPA provides no positive or negative candidates, so tissue controls cannot be selected from this evidence.
HPA tissue expression evidence for MAK
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 MAK Western Blot Tips
Deeper troubleshooting and optimisation questions for MAK, answered from its protein features.
How should MAK 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 MAK isoforms produce different bands?
Isoforms · UniProt lists three isoforms. Relative to isoform 1, isoform 2 replaces A at position 532 with AEESIIKPIEKLSCNETFPEKLEDPQ, while isoform 3 lacks residues 533–572. These sequence differences could affect apparent size, but the features do not establish which isoforms produce visible bands.
For a shared MAK signal, choose an epitope outside the sequence differences at position 532 and residues 533–572. An epitope within those regions may recognize isoforms differently. Check the antibody’s stated epitope before interpreting relative band intensities.
For a shared MAK signal, choose an epitope outside the sequence differences at position 532 and residues 533–572. An epitope within those regions may recognize isoforms differently. Check the antibody’s stated epitope before interpreting relative band intensities.
Which MAK phosphorylation sites matter when interpreting a band shift?
PTM · UniProt annotates phosphothreonine at position 157 and phosphotyrosine at position 159, both by autocatalysis. A phosphorylation-sensitive assay could help assess modification, but these sites alone do not establish a visible shift. These are UniProt coordinates; antibody or paper numbering may differ.
Does this guide establish induction of MAK?
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 MAK?
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 A00407-1 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 MAK be quantified?
Quantitation · Standard workflow guidance: quantify only a validated target band within the linear exposure range. Use consistent sample preparation and loading, retain biological replicates, and avoid interpreting saturation or loading differences as regulation.
How does MAK’s observed band compare with its predicted mass?
Interpretation · MAK’s predicted mass is 70.6 kDa, and the supplied observed band is approximately 70 kDa. The annotated features do not establish a reason for any small difference in apparent mobility.
MAK is annotated in the nucleus and at cytoskeletal structures, including centrosomes, spindles, and the midbody; ciliary and photoreceptor locations are also listed. Keep sample preparation and fraction selection consistent when comparing signals, since the sampled compartments can affect how much MAK is recovered.
Compare them with the approximately 70 kDa observed band and consider the three annotated isoforms, particularly the insertion in isoform 2 and deletion in isoform 3. The phosphorylation sites at UniProt positions 157 and 159 do not by themselves identify an unexpected band or prove a mobility shift.
MAK is annotated in the nucleus and at cytoskeletal structures, including centrosomes, spindles, and the midbody; ciliary and photoreceptor locations are also listed. Keep sample preparation and fraction selection consistent when comparing signals, since the sampled compartments can affect how much MAK is recovered.
Compare them with the approximately 70 kDa observed band and consider the three annotated isoforms, particularly the insertion in isoform 2 and deletion in isoform 3. The phosphorylation sites at UniProt positions 157 and 159 do not by themselves identify an unexpected band or prove a mobility shift.
Boster reagents
MAK 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-MAK Antibody Picoband®
Cat # A00407-1
The catalog reports one anti-MAK antibody, A00407-1, with a Western blot image showing an approximately 70 kDa band in human HepG2 and MCF-7 whole cell lysates. The supplied evidence does not include independent validation.
Which to pick: A00407-1 is the only listed option. It is marked reactive with human samples and has a Western blot image from HepG2 and MCF-7 lysates; suitability for other sample types is not established here.
Source: BosterBio MAK gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P20794.
- Human Protein Atlas. MAK tissue expression.
- PMC3820702 — target-verified WB comparison
- PMC11412320 — target-verified WB comparison