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ADAM19 · Western blot design guide
Design a Western Blot for ADAM19
Real validated ADAM19 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-ADAM19 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Last reviewed: May 2026 · Scientific review: Boster Bio technical team
ADAM19 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 | ~105 kDa | |
| Observed band | ~105 kDa | |
| Gel | 8–10% | |
| Negative control | siRNA / KO lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | N-glycosylation increases apparent size | |
| Regulation | Il2 stat5 signaling | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated ADAM19 Western Blot Protocols
Literature-validated Western blot parameters for ADAM19 — gel percentage, transfer, blocking, antibody incubation and detection, extracted from published methods.
| Sample / lysate | human U251 , Lane 2: human SH-SY5Y , Lane 3: human U87 , Lane 4: human U20S . After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA 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-ADAM19 antigen affinity purified polyclonal antibody (Catalog # PA2070) 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 1:5000 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 ADAM19 at approximately 105 kDa. The expected band size for ADAM19 is at 105 kDa |
| Gel % | 8–10% |
| Load | 30 ug |
| 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, 1:5000 |
| Wash | TBS-0.1% Tween, 3 × 5 min |
| Detection | ECL |
| Exposure / imaging | Tanon 5200 |
| Observed band | 105 kDa |
Section 2
What Is the Expected ADAM19 Western Blot Band Size?
ADAM19 runs at its ~105 kDa predicted mass on blots because glycosylation at four N-linked sites offsets the mass lost from signal-peptide and propeptide cleavage during maturation.
| single band at approximately 105 kDa | corresponds to the mature, fully N-glycosylated, membrane-anchored form of ADAM19 seen in most whole-cell lysates |
| diffuse or smeared signal around the ~105 kDa region rather than a crisp line | heterogeneous occupancy of the four N-linked glycosylation sites (Asn144, Asn444, Asn447, Asn645) broadens the apparent band |
| an extra, higher band above the mature ~105 kDa species | residual uncleaved zymogen still carrying the inhibitory propeptide (residues 26-202) that has not yet been processed to the mature protease |
| a band noticeably lower than ~105 kDa, closer to an unmodified core protein | cleavage of the signal peptide (1-25) and propeptide (26-202) without full compensating glycosylation, giving a smaller mature backbone |
| two closely spaced bands (doublet) rather than one | alternative splicing generates isoforms A and B that differ slightly in length and migrate at slightly different apparent sizes |
💡Expected ADAM19 appearanceExpect a single band at approximately 105 kDa, representing mature, N-glycosylated ADAM19 after signal-peptide and propeptide cleavage, with glycosylation at four N-linked sites largely offsetting the mass lost during propeptide removal.
| Predicted mass of the full-length precursor (UniProt) | gives a 105 kDa baseline calculated from the unmodified 955-aa sequence before any processing or glycosylation is considered |
| N-glycosylation at Asn144, Asn444, Asn447, and Asn645 | adds variable carbohydrate mass, shifting and broadening the apparent band and largely explaining the ~105 kDa observed size |
| Signal peptide (residues 1-25) and propeptide (residues 26-202) cleavage | removes roughly 20+ kDa from the precursor to yield the smaller, catalytically active mature enzyme before glycosylation adds mass back |
| Alternative splicing into isoforms A and B | produces two protein forms of differing length that can resolve as separate bands of slightly different apparent size |
| Reducing vs non-reducing conditions with seven intra-chain disulfide bonds | incomplete reduction leaves the protein more compact and disulfide-constrained, altering its migration relative to the fully reduced form |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | ADAM19 is a single-pass type I membrane protein that stays cell-associated and partitions into membrane fractions if lysis is too mild | use a detergent-based lysis buffer (e.g., RIPA or NP-40 based) that solubilizes membrane proteins and include membrane fractions in the prep |
| Band higher than expected | incomplete conversion of the zymogen, leaving propeptide-containing precursor alongside heavily glycosylated mature protein | compare band position to the ~105 kDa mature species and check for a distinct higher precursor band consistent with unprocessed propeptide |
| Band lower than expected | mature ADAM19 after signal-peptide and propeptide removal runs below 105 kDa when glycosylation is reduced or absent, as with non-glycosylated recombinant standards | treat a parallel sample with PNGase F or compare to a mammalian-expressed standard to confirm the contribution of glycosylation to the ~105 kDa size |
| Broad smear instead of sharp band | heterogeneous occupancy and processing of the four N-linked glycosylation sites produces a range of glycoforms | deglycosylate the sample enzymatically to collapse the smear into a single sharper core-protein band for comparison |
| Multiple bands | coexistence of isoforms A and B, or of unprocessed zymogen alongside mature cleaved protein, in the same lysate | map detected bands against the expected precursor, mature, and isoform sizes and confirm the antibody epitope is present in the relevant region |
| Fragments below expected size | ADAM19 is itself a metalloprotease and can undergo self- or co-incubated proteolytic cleavage during sample handling | include protease inhibitors during lysis and minimize processing time to limit degradation before running the gel |
Experiment evidence
Western blot images showing ADAM19 band
PA2070 · ADAM19
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for ADAM19 Western blot
🧪For positive controls for ADAM19 in Western blot, you can use lysate from cells engineered to overexpress ADAM19, since no tissue or cell line expression data is available in the Human Protein Atlas for this protein.
Positive control: ADAM19-overexpressing cell lysate
Negative control: no HPA expression data; use ADAM19 siRNA knockdown or KO cell line
Loading controls: Run GAPDH, β-actin, and a total-protein stain (e.g., stain-free gel, Ponceau S, or REVERT) as loading controls.
⚠️Feasibility: As a single-pass type I membrane protein with no HPA expression profiling available, endogenous tissue-based controls cannot be selected with confidence, so an overexpression system paired with siRNA/KO validation is the more feasible approach.
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 |
|---|
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|
Section 3
Advanced ADAM19 Western Blot Tips
Deeper troubleshooting and optimisation questions for ADAM19, answered from its protein features.
Why does ADAM19 run close to its predicted 105 kDa mass?
ADAM19 is synthesized as a zymogen; UniProt predicts 105 kDa for the full precursor, matching the observed ~105 kDa band. The propeptide (residues 26-202) is removed during activation, so a mature processed form would run several kDa lower. Detecting the band near 105 kDa indicates the full-length precursor is the dominant species in the sample.
Could isoforms A and B appear as separate bands?
ADAM19 has two annotated isoforms (A and B) arising from alternative splicing. Differential exon inclusion can alter length and apparent molecular weight, so tissues expressing both isoforms may show a doublet near 105 kDa rather than a single band. Isoform-specific antibodies or peptide mapping can confirm which variant is detected.
Does zymogen activation change the detected band pattern?
ADAM19 is produced as an inactive zymogen with an N-terminal propeptide spanning residues 26-202. Proprotein convertases remove this propeptide to generate the active, membrane-anchored protease. Because both unprocessed precursor and processed mature protein can coexist in a lysate, blots may show the full-length ~105 kDa precursor alongside a lower molecular weight mature band.
Which blocking buffer minimizes background for ADAM19?
ADAM19 carries four predicted N-glycosylation sites, and glycoproteins in milk-based blockers can cause lectin-like cross-reactivity and background smearing. Blocking with 5% BSA in TBST for one hour typically gives cleaner backgrounds than milk for this glycosylated membrane protein.
What transfer method to use for ADAM19 Western blot?
ADAM19 is a ~105 kDa single-pass type I membrane glycoprotein stabilized by seven disulfide bonds. Reduce samples with DTT or beta-mercaptoethanol before SDS-PAGE to linearize the protein, then use wet/tank transfer with extended transfer time to efficiently move this high molecular weight protein onto PVDF membrane.
How to normalize ADAM19 signal for quantitation?
Because ADAM19 is membrane-anchored, extraction efficiency and loading can vary between membrane and cytosolic fractions, making cytosolic housekeeping proteins unreliable normalizers. Use a total protein stain such as Ponceau S or REVERT on the same membrane to normalize ADAM19 signal for accurate relative quantitation.
What causes extra bands besides the 105 kDa band?
Extra bands can arise from variable occupancy of the four N-glycosylation sites, producing smeared or shifted species. Non-reducing conditions may preserve disulfide-linked (seven bonds) aggregates at higher molecular weight. A lower band can reflect the mature protease after propeptide (26-202) removal, distinct from the full-length zymogen precursor.
Boster reagents
Best ADAM19 Western Blot Antibodies
BosterBio's ADAM19 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-ADAM19 Antibody Picoband®
Cat # PA2070
Our recommended anti-ADAM19 Western blot antibodies are top-performing and extensively cited, having undergone rigorous validation including orthogonal cross-checks against negative tissue controls and complementary detection methods, ensuring specific, reproducible ADAM19 detection you can rely on for confident results.
Which to pick: Only one Boster antibody is listed for ADAM19: PA2070, which includes an actual Western blot validation image demonstrating its performance—making it the clear, straightforward choice for your ADAM19 WB experiments.
Source: BosterBio ADAM19 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q9H013.
- Human Protein Atlas. ADAM19 tissue expression.