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- Table of Contents
MDH1 / Malate dehydrogenase, cytoplasmic · Western blot design guide
Design a Western Blot for MDH1
Real validated MDH1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-MDH1 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
MDH1 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 | ~36.4 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Cerebral cortex (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 | Phosphorylated + Acetylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 3 isoform(s) |
Section 1
Real Curated MDH1 Western Blot Protocols
The A04262 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | Hela, HepG2, PC12 (catalog A04262) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A04262; 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 MDH1 Western Blot Band Size?
MDH1 has a predicted 36.4 kDa monomer; isoforms and homodimerization could affect patterns, but altered migration is not demonstrated.
| Band near 36.4 kDa | Consistent with the predicted MDH1 monomer; confirm identity with antibody controls |
| Band near 73 kDa | Could reflect retained MDH1 homodimer, although its persistence during electrophoresis is unestablished |
| Several discrete bands | Could include MDH1 isoforms 1, 2, and 3; their migration differences are unknown |
| Single band away from 36.4 kDa | Could represent an MDH1 isoform, but its identity requires confirmation |
💡Expected MDH1 appearanceThe predicted MDH1 monomer is 36.4 kDa; no empirical band size is supplied, and any additional band requires identity controls because isoform migration and dimer retention are unestablished.
| Predicted MDH1 mass | Places the predicted monomer near 36.4 kDa |
| Homodimer formation | Could yield a band near twice the monomer mass if the dimer survives preparation and electrophoresis |
| Isoform 1 | May migrate differently from isoforms 2 and 3; relative size is unknown |
| Isoform 2 | May migrate differently from isoforms 1 and 3; relative size is unknown |
| Isoform 3 | May migrate differently from isoforms 1 and 2; relative size is unknown |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | MDH1 homodimer may persist, or the band may be unrelated | Compare fully denaturing conditions and confirm antibody specificity |
| Band lower than expected | An MDH1 isoform is possible, but its mass is not supplied | Confirm the band with an independent MDH1 antibody or MDH1 depletion |
| Multiple bands | MDH1 has three named isoforms, but distinct migration is unestablished | Compare bands after MDH1 depletion and check antibody specificity |
| No band in lysate | Cytosolic MDH1 may be below detection in the loaded sample | Check lysate loading and use a positive control lysate |
| Weak or no signal | Insufficient detectable cytosolic MDH1 or antibody sensitivity | Increase lysate input and verify the antibody with a positive control |
Experiment evidence
Western blot images showing MDH1 band
A04262 · MDH1
A04262-2 · MDH1
A04262-1 · MDH1
M04262 · MDH1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for MDH1 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for MDH1 in Western blot, you can use cerebral cortex tissue, which HPA rates High.
Positive control: Cerebral cortex (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: Cytosolic MDH1 should be accessible in tissue lysate, though HPA non-detection in adipose tissue may not guarantee a blank blot.
HPA tissue expression evidence for MDH1
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 |
|---|---|---|---|---|
| Cerebral cortex | neuropil | High | Protein (IHC) | HPA → |
| Stomach | glandular cells | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Caudate | neuronal cells | Medium | Protein (IHC) | HPA → |
| Cerebellum | Purkinje 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 → |
| Appendix | glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | adipocytes | Not detected | Protein (IHC) | HPA → |
| Cervix | glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced MDH1 Western Blot Tips
Deeper troubleshooting and optimisation questions for MDH1, answered from its protein features.
Should an MDH1 band appear exactly at 36.4 kDa?
Band shift · 36.4 kDa is the predicted mass of canonical MDH1; no observed band position is supplied. The listed modifications alone do not establish a visible shift or explain any difference between predicted and apparent mass.
How might MDH1 isoforms affect band interpretation?
Isoforms · UniProt lists isoforms 1, 2, and 3. Isoform 2 lacks canonical residues 1–89, while isoform 3 replaces canonical residue 1 with MRRCSYFPKDVTVFDKDDK. These sequence differences could affect antibody recognition or migration; the supplied features do not establish their observed band positions.
For detection of isoform 2, choose an epitope outside canonical residues 1–89, which it lacks. Isoform 3 has a different N terminus. Check the antibody’s stated epitope against each sequence before interpreting a missing band as absent MDH1.
For detection of isoform 2, choose an epitope outside canonical residues 1–89, which it lacks. Isoform 3 has a different N terminus. Check the antibody’s stated epitope against each sequence before interpreting a missing band as absent MDH1.
Which MDH1 modifications should I consider when interpreting bands?
PTM · UniProt lists N-acetylserine 2; succinyllysine 110, 214, 298, and 318; acetyllysine 118, 121, and 298; phosphoserine 217, 241, 309, 332, and 333; and methylarginine 230. These are canonical UniProt coordinates, which may differ from paper or antibody numbering. Their presence does not establish a visible band shift.
UniProt lists N6-acetyllysine and N6-succinyllysine as alternate modifications at canonical position 298. Interpret them as alternatives at that residue, not simultaneous modifications on one molecule. Neither annotation predicts a distinct Western blot band.
UniProt lists N6-acetyllysine and N6-succinyllysine as alternate modifications at canonical position 298. Interpret them as alternatives at that residue, not simultaneous modifications on one molecule. Neither annotation predicts a distinct Western blot band.
Does this guide establish induction of MDH1?
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 MDH1?
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 A04262 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 I quantify MDH1 if multiple bands appear?
Quantitation · Decide whether the measurement targets all detected MDH1 or a specific isoform, then quantify only bands supported by antibody recognition and appropriate band identification. Isoforms 2 and 3 differ in sequence from canonical MDH1, and the supplied features provide no observed band positions. Avoid assigning bands by size alone.
Could an unexpected MDH1 band represent a dimer?
Interpretation · UniProt describes MDH1 as a cytosolic homodimer, but that annotation alone does not identify an unexpected Western blot band as a dimer. Consider the three isoforms and antibody epitope when assessing additional bands; confirm their identity before assigning them to MDH1.
Boster reagents
MDH1 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-MDH1 Antibody
Cat # A04262
Real WB data
Anti-MDH1 Antibody Picoband®
Cat # A04262-2
Real WB data
Anti-MDH1 Antibody (C-term)
Cat # A04262-1
Real WB data
Anti-MDH1 Rabbit Monoclonal Antibody
Cat # M04262
All four listed anti-MDH1 antibodies have WB images. A04262-2 shows human cell and rat and mouse tissue lysates, but its caption reports a band near 36 kDa against an expected 59 kDa. Check band identity in your samples.
Which to pick: For human cell or rodent tissue lysates, A04262-2 has directly pictured examples. A04262 shows HeLa, HepG2, and PC12 lysates; M04262 shows HeLa. A04262-1 lists human reactivity and shows HL-60 and THP-1 lysates.
Source: BosterBio MDH1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P40925.
- Human Protein Atlas. MDH1 tissue expression.
- PMC10392054 — target-verified WB comparison
- PMC12116433 — target-verified WB comparison
- PMC5501748 — target-verified WB comparison