This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
MST1 · Western blot design guide
Design a Western Blot for MST1
Real validated MST1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-MST1 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Last reviewed: May 2026 · Scientific review: Boster Bio technical team
MST1 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 | ~80.3 kDa | |
| Observed band | ~80 kDa | |
| Gel | 8–10% | |
| Positive control | Adipose tissue+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Alpha/beta chain cleavage | |
| Regulation | Coagulation | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated MST1 Western Blot Protocols
Literature-validated Western blot parameters for MST1 — gel percentage, transfer, blocking, antibody incubation and detection, extracted from published methods.
| Sample / lysate | human Hela , Lane 2: human MCF-7 , Lane 3: human HepG2 , Lane 4: human SK-OV-3 , Lane 5: human A549 , Lane 6: rat kidney , Lane 7: mouse kidney . After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-MST1 antigen affinity purified polyclonal antibody (Catalog # A01069-1) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for MST1 at approximately 80KD. The expected band size for MST1 is at 80KD |
| Gel % | 8–10% |
| Load | 50ug |
| Transfer | nitrocellulose membrane, 150 mA, 50–90 min |
| Membrane | nitrocellulose |
| Blocking | 5% non-fat milk / TBS, 1.5 h RT |
| Primary antibody | 0.5 µg/mL |
| Primary incubation | overnight at 4 °C |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:10000 |
| Wash | TBS-0.1% Tween, 3 × 5 min |
| Detection | ECL |
| Exposure / imaging | Tanon 5200 |
| Observed band | 80 kDa |
Section 2
What Is the Expected MST1 Western Blot Band Size?
MST1 has an 80.3 kDa predicted core but is glycosylated and disulfide-linked into an alpha/beta heterodimer, matching the empirically observed ~80 kDa band.
| single band at approximately 80 kDa | matches the glycosylated single-chain pro-MST1 precursor, the dominant form detected in cells and serum |
| band running slightly above the 80.3 kDa predicted mass, sometimes smeared | N-linked glycosylation at Asn72, Asn296, and Asn615 adds carbohydrate mass and microheterogeneity |
| band near 80 kDa retained intact under non-reducing conditions | the proteolytically generated alpha and beta chains stay joined by an inter-chain disulfide bond, co-migrating as one complex |
| loss of the ~80 kDa band and appearance of smaller chains after strong reduction | reducing agents break the inter-chain disulfide, separating the disulfide-linked alpha/beta heterodimer into its two component chains |
| little or no band in whole-cell lysate | MST1 is a secreted protein, so most of the mature protein leaves the cell rather than accumulating intracellularly |
💡Expected MST1 appearanceExpect a single band at approximately 80 kDa, matching the glycosylated single-chain pro-MST1; the disulfide-linked alpha/beta heterodimer formed after processing co-migrates near this same 80 kDa size unless reduced.
| Predicted mass (80.3 kDa, UniProt) | sets the baseline single-chain polypeptide size before any modification |
| N-glycosylation at Asn72, Asn296, and Asn615 | adds carbohydrate mass and heterogeneity, nudging the band above the 80.3 kDa core and broadening it |
| Inter-chain disulfide linking the alpha and beta chains | holds the proteolytically processed chains together near 80 kDa under non-reducing conditions; full reduction resolves them into separate smaller alpha and beta chains |
| Signal peptide (residues 1-18) | is cleaved during secretion, so the mature secreted protein runs slightly smaller than the unprocessed translated precursor |
| Secreted subcellular location | shifts most mature protein into conditioned media or serum, lowering the signal detected in whole-cell lysate |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | MST1 is secreted, so little full-length or processed protein remains intracellular | probe concentrated conditioned media or serum/plasma instead of, or in addition to, whole-cell lysate |
| Band higher than expected | N-glycosylation at three sites (Asn72, Asn296, Asn615) adds mass above the 80.3 kDa predicted core | treat lysate with PNGase F and confirm the band shifts down toward the predicted mass |
| Band lower than expected | cleavage of the signal peptide during secretion yields a mature protein smaller than the full translated precursor | compare against a construct or standard known to retain versus lack the signal peptide before concluding degradation |
| Broad smear instead of sharp band | heterogeneous occupancy and processing of the three N-glycosylation sites broadens the apparent mass | run a lower-percentage or longer gel, or deglycosylate with PNGase F for a tighter band |
| Multiple bands | partial versus complete reduction of the inter-chain disulfide linking the alpha and beta chains yields both the intact ~80 kDa complex and separated chains | standardize on either fully reducing or fully non-reducing sample buffer across lanes for consistent comparison |
| Fragments below expected size | proteolytic processing of the alpha/beta chains during endogenous maturation or sample handling can generate smaller species | add protease inhibitors during lysis, keep samples cold, and avoid repeated freeze-thaw |
Experiment evidence
Western blot images showing MST1 band
A01069-1 · MST1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for MST1 Western blot
🧪For positive controls for MST1 in Western blot, you can use human serum or hepatocyte-conditioned culture medium, since the supplied HPA tissue panel shows no clear positive lysate signal for this secreted, hepatocyte-associated protein.
Positive control: Serum or conditioned medium
Negative control: Adipose tissue
Loading controls: Include GAPDH and β-actin alongside a total-protein stain (e.g., stain-free gel, Ponceau S, or REVERT) as loading controls.
⚠️Feasibility: Being a secreted protein, MST1 can give weak or absent signal in whole-cell/tissue lysates, so conditioned medium or serum/plasma samples give more reliable positive controls than standard lysate blots.
HPA tissue expression evidence for MST1
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.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | 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 → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | 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 → |
Section 3
Advanced MST1 Western Blot Tips
Deeper troubleshooting and optimisation questions for MST1, answered from its protein features.
Why does MST1 run near its predicted 80 kDa mass?
MST1 is secreted with a cleaved 18-residue signal peptide and is glycosylated, but its predicted 80.3 kDa mass already matches the reported ~80 kDa band, so little correction from the sequence-based prediction is needed when run under non-reducing SDS-PAGE conditions that preserve the intact disulfide-linked chain.
Why might MST1 show two bands on reducing gels?
MST1 is a dimer of an alpha chain and a beta chain joined by a disulfide bond. Reducing agents like DTT or BME break this bond, splitting the ~80 kDa full-length species into separate lower-molecular-weight alpha- and beta-chain bands. Run parallel reducing and non-reducing lanes to confirm chain composition.
Could N-glycosylation shift or smear the MST1 band?
MST1 has 3 predicted N-glycosylation sites. Heterogeneous glycan occupancy across these sites can broaden or slightly upshift the band relative to the unmodified polypeptide mass. If smearing is observed, PNGase F treatment before loading can deglycosylate the sample and sharpen the resulting band.
What transfer method to use for MST1 Western blot?
Given MST1's ~80 kDa size and 20 disulfide bonds, use wet (tank) transfer with a low-percentage acrylamide gel and an extended transfer time to efficiently elute the large, compact, disulfide-stabilized protein. Including SDS in the transfer buffer can further aid its solubilization and movement onto the membrane.
How should blocking be optimized for glycosylated MST1?
Because MST1 carries 3 N-glycans, glycoprotein-rich blockers such as non-fat milk can occasionally elevate background through carbohydrate-mediated cross-reactivity. A BSA-based blocking buffer is a safer default for clean detection of this glycoprotein, especially if lectin- or glycan-sensitive detection reagents are used downstream.
How should MST1 levels be quantified reliably?
As a secreted protein with no cytoplasmic retention, MST1 is best quantified from conditioned media, serum, or plasma rather than whole-cell lysates. Normalize loading to total protein stain, such as Ponceau S, rather than to intracellular housekeeping proteins that are not co-secreted with MST1.
What explains unexpected extra bands on an MST1 blot?
Extra bands can reflect incompletely reduced disulfide-linked dimer, separated alpha and beta chains under reducing conditions, or heterogeneous occupancy across the 3 glycosylation sites. Comparing reducing versus non-reducing runs, and including a deglycosylation control, helps distinguish these expected chain forms from genuine nonspecific bands.
Boster reagents
Best MST1 Western Blot Antibodies
BosterBio's MST1 antibodies are among the best-performing WB antibodies on the market — well cited, thoroughly validated, and orthogonally cross-validated against negative tissues and complementary methods.
Real WB data
Anti-MST1 Antibody Picoband®
Cat # A01069-1
Boster's anti-MST1 antibodies are top-performing, widely cited reagents rigorously validated for western blot specificity, cross-confirmed against negative-tissue controls and orthogonal detection methods to ensure confident, reproducible MST1 detection.
Which to pick: Only one Boster anti-MST1 antibody is listed, A01069-1, which includes an actual western blot validation image—making it the clear, straightforward choice for your MST1 WB experiments.
Source: BosterBio MST1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P26927.
- Human Protein Atlas. MST1 tissue expression.