15% OFF arrow_drop_up
Skip to content

Join our rewards club and start earning on every purchase today

Get in touch with us

4–20% Gradient Gels vs. Fixed % Gels: When to Use Each

When you’re prepping for SDS-PAGE, selecting the right gel is more than just grabbing whatever’s available — it directly affects how well your proteins separate and whether you can even see your target bands. This article breaks down the core differences between fixed % acrylamide gels and 4–20% gradient gels, so you can confidently choose the best format for your experiment. Whether you're analyzing large complexes, small peptides, or multiple targets in one run, the right gel will save time and give you sharper, more reproducible results.

 Fixed % Gels

  • Made with one acrylamide concentration throughout (e.g. 10%, 12%, or 15%)

  • Pore size is uniform across the gel

  • Best for resolving proteins within a narrow molecular weight range

Gradient Gels (e.g. 4–20%)

  • Acrylamide concentration increases gradually from the top (stacking zone) to the bottom (resolving zone)

  • Pore size narrows from top to bottom

  • Enables separation of a wide range of protein sizes in a single run


When to Use Fixed % Gels

Use fixed % gels when:

  • You know your target protein's approximate size

  • You want maximum resolution around a specific molecular weight

  • You're running similar-sized proteins across multiple lanes

Examples:

Gel % Protein Range Best For
10% 50–150 kDa Actin, tubulin, HSP70
12% 20–100 kDa Enzymes, transcription factors
15% <30 kDa

Ubiquitin, small peptides

 

🎯 Tip: Fixed % gels are ideal for labs that focus on 1–2 known targets and require sharp, clean bands in that range.

🧪 When to Use 4–20% Gradient Gels

Use gradient gels when:

  • Your protein mixture contains multiple sizes

  • You're working with unknown molecular weights

  • You want to visualize full molecular weight ladders

  • You're analyzing post-translational modifications or protein degradation products

Why Gradient Gels Are Versatile:

  • Small proteins resolve at the top (low acrylamide %)

  • Large proteins resolve at the bottom (high acrylamide %)

  • This allows high and low MW bands to separate cleanly on one gel

🧪 Use case: Western blots where you want to probe both a phosphorylated 25 kDa target and its full-length 80 kDa form — a gradient gel is perfect.


⚖️ Comparison Table: Gradient vs. Fixed % Gels

Feature Fixed % Gel 4–20% Gradient Gel
Pore Size Uniform Varies (top to bottom)
Resolution Range Narrow Wide
Best For Known MW proteins Mixed/unknown sizes
Run Time Slightly faster Slightly longer
Price Lower Slightly higher
Reproducibility High High (precast)
Visualization Sharp bands at specific MW Ladder-like separation across range



🧪 NuSep’s Precast Gel Options

NuSep offers a wide range of fixed % gels (10%, 12%, 15%) and 8-16%, 4–20% gradient gels across three compatibility formats:

  • NB Series – Bio-Rad® Mini-PROTEAN® tanks

  • NG Series – Cytiva™ / Hoefer® systems

  • NN Series – Invitrogen™ Novex® tanks

✅ All gels are leak-resistant, shelf-stable, and come with tight well formation for clean, reproducible results.

👉 Explore NuSep Precast Gels


❓FAQs

Q: Do gradient gels require a special tank?
A: No — as long as the gel cassette fits your tank (e.g. Mini-Pro, Bio-Rad, Hoefer, Invitrogen), gradient and fixed % gels run the same way.

Q: Can I cut gradient gels for transfer?
A: Yes, just be mindful that protein migration rates vary across the gel — always use molecular weight markers.

Q: Do gradient gels affect blotting efficiency?
A: Slightly — very large proteins may transfer more slowly from dense (high %) zones. Use extended transfer times if needed.