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DLL1 · Western blot design guide
Design a Western Blot for DLL1
Real validated DLL1 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-DLL1 WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Last reviewed: May 2026 · Scientific review: Boster Bio technical team
DLL1 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 | ~78.1 kDa | |
| Gel | 10–12% | |
| Negative control | siRNA / KO lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Phosphorylated | |
| Caveat | N-glycosylation increases mass | |
| Regulation | Notch signaling | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated DLL1 Western Blot Protocols
Literature-validated Western blot parameters for DLL1 — gel percentage, transfer, blocking, antibody incubation and detection, extracted from published methods.
| Sample / lysate | human HELA , Lane 2: monkey COS-7 , Lane 3: huamn Jurkat , Lane 4: huamn Raji , Lane 5: human SW620 , Lane 6: human HepG2 , Lane 7: human U20S , Lane 8: rat heart , Lane 9: rat C6 , Lane 10: mouse heart , Lane 11: mouse Neuro-2a , Lane 12: mouse RAW264.7 , 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-DLL1 antigen affinity purified polyclonal antibody (Catalog # A02513-1) at 0.25 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 DLL1 at approximately 78KD. The expected band size for DLL1 is at 78KD |
| Gel % | 10–12% |
| Load | 50ug |
| Transfer | nitrocellulose membrane, 150 mA, 50–90 min |
| Membrane | nitrocellulose |
| Blocking | 5% non-fat milk / TBS, 1.5 h RT |
| Primary antibody | 0.25 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 | 78 kDa |
Section 2
What Is the Expected DLL1 Western Blot Band Size?
DLL1 has a 78 kDa predicted backbone but likely runs somewhat higher due to N-glycosylation at Asn477, with a possible dimer band if reduction is incomplete.
| single band running a bit above 78 kDa | N-linked glycosylation at Asn477 adds carbohydrate mass to the mature single-pass membrane glycoprotein |
| band at or slightly below the 78 kDa unmodified backbone | cleavage of the 17-residue signal peptide removes the leader sequence from the precursor during membrane insertion |
| faint doublet or extra band near the main monomer band | DLL1 has two annotated splice isoforms that can resolve as distinct, closely spaced species |
| higher-molecular-weight band near double the monomer size | DLL1 is annotated as a homodimer, so incompletely reduced or non-reducing samples can retain dimeric complexes |
| reasonably sharp band with only mild broadening, not a heavy smear | a single N-glycosylation site limits glycoform heterogeneity compared with heavily glycosylated proteins |
💡Expected DLL1 appearanceUnder reducing SDS-PAGE, expect a single monomer band running somewhat above the 78 kDa predicted mass, reflecting N-glycosylation at Asn477 on the signal-peptide-cleaved mature protein, with a fainter higher band if dimer persists.
| Predicted mass (78.1 kDa, 723 aa) | sets the baseline unmodified backbone size before any post-translational changes |
| N-glycosylation at Asn477 | adds carbohydrate mass, shifting the mature band above the 78 kDa backbone and adding modest heterogeneity |
| Signal peptide cleavage (residues 1-17) | removes the N-terminal leader sequence so the mature membrane-inserted form runs slightly smaller than the unprocessed precursor |
| Homodimer formation | can yield a higher-order species near twice the monomer mass if dimers survive non-reducing or incompletely denaturing conditions |
| Splice isoforms (1 and 2) | different isoforms can shift the apparent monomer position or add a secondary band, though relative sizes are only qualitative |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | incomplete reduction leaves homodimer complexes intact, pushing the band toward roughly double the monomer mass | increase reducing agent concentration and heating time before loading to fully dissociate dimers |
| Broad smear instead of sharp band | heterogeneous glycosylation at the single N-linked site broadens the band envelope | treat lysate with PNGase F or resolve on a lower-percentage gel to sharpen the glycoform spread |
| Multiple bands | the two annotated splice isoforms of DLL1 can appear as distinct, closely sized species | compare against a known isoform-expressing control and check isoform-specific literature for the cell/tissue type |
| Band lower than expected | signal peptide cleavage removes the N-terminal leader and full reduction fully dissociates any dimer, leaving the mature monomer near or below the unmodified predicted mass | confirm against a positive control lysate and check that reducing conditions are not causing additional degradation |
| Weak or no signal | as an apical membrane, adherens-junction, and membrane-raft-associated protein, standard aqueous lysis buffers may not efficiently solubilize DLL1 | use a detergent-based membrane lysis buffer and increase total protein loaded per lane |
Experiment evidence
Western blot images showing DLL1 band
A02513-1 · DLL1
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for DLL1 Western blot
🧪For positive controls for DLL1 in Western blot, you can use lysate from cells engineered to overexpress DLL1, such as transiently transfected HEK293 cells, since no Human Protein Atlas tissue expression data are available for this protein.
Positive control: DLL1-transfected HEK293 cells
Negative control: ubiquitously expressed; use siRNA knockdown or KO line
Loading controls: Run GAPDH and β-actin antibody blots alongside a total-protein stain (e.g., stain-free gel imaging, Ponceau S, or REVERT) to confirm equal loading.
⚠️Feasibility: As a single-pass membrane/adherens-junction glycoprotein with no HPA expression data to guide tissue selection, clean controls are hard to source, so an overexpression system paired with siRNA/KO validation is the most reliable 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 DLL1 Western Blot Tips
Deeper troubleshooting and optimisation questions for DLL1, answered from its protein features.
Why might DLL1 run higher than its predicted 78 kDa mass?
N-glycosylation at one site plus the extracellular EGF-repeat domain's dense disulfide bonding can retard SDS-PAGE mobility, so DLL1 often migrates near 85-100 kDa rather than the calculated 78.1 kDa mass.
Could the two DLL1 isoforms appear as separate bands?
DLL1 has two annotated isoforms from alternative splicing; if your antibody epitope is shared, expect a faint secondary band at a different apparent mass alongside the canonical full-length product.
Is DLL1 phosphorylation likely to affect blot pattern?
DLL1 carries two modified residues consistent with phosphorylation. Phosphorylated pools can shift mobility slightly or appear as a doublet; Notch pathway activation state in your cell model may change the ratio.
What blocking buffer avoids interference with DLL1 glycosylation?
Since DLL1 has an N-glycosylation site, avoid milk-based blockers, which contain glycoproteins/lectins that raise background on glycosylated targets; use BSA-based blocking instead.
What transfer method to use for DLL1 Western blot?
DLL1 is a large (78 kDa), disulfide-rich (27 bonds), single-pass type I membrane glycoprotein; use wet transfer with PVDF membrane and adequate SDS/methanol balance to ensure complete transfer of the reduced monomer.
Should samples be run under reducing conditions for DLL1?
Yes. DLL1 forms homodimers and contains 27 disulfide bonds in its extracellular EGF repeats; without full reduction (DTT or beta-mercaptoethanol), it can run at a higher apparent mass or as aggregates, complicating accurate quantitation.
Why might an unexpected higher-MW DLL1 band appear?
DLL1 undergoes isopeptide (ubiquitin-like) conjugation and forms complexes with CTNNB1, CDH2, and MAGI1 at adherens junctions; incomplete lysis or reduction can retain these interactions, producing higher-molecular-weight bands beyond the monomer.
Boster reagents
Best DLL1 Western Blot Antibodies
BosterBio's DLL1 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-DLL1 Antibody Picoband®
Cat # A02513-1
Our recommended anti-DLL1 antibody is a best-performing, extensively cited reagent, rigorously validated for Western blot specificity and orthogonally cross-validated against negative tissue controls and complementary detection methods, ensuring confident, reproducible detection of DLL1 protein across experimental systems.
Which to pick: Only one Boster anti-DLL1 antibody is catalogued here, A02513-1, which includes an actual Western blot validation image, making it the clear, and only, choice for DLL1 WB experiments in this guide.
Source: BosterBio DLL1 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry O00548.
- Human Protein Atlas. DLL1 tissue expression.