High CV from Bubbles in Wells

Symptom

Inconsistent absorbance readings between replicate wells with coefficient of variation >15%. Visual inspection may reveal bubbles present in wells, particularly when using detergent-containing solutions.

1. 1 Solutions containing detergents are especially susceptible to bubble formation
2. 2 Rapid or forceful addition of reagents to wells introduces air
3. 3 Bubbles remaining in wells interfere with optical reading path
4. 4 Multi-channel pipettors may trap air when dispensing detergent solutions

1. 1 Ensure no bubbles are present prior to reading the plate by visual inspection
2. 2 Gently add solutions to the wells to minimize turbulence and air incorporation
3. 3 Pop bubbles by touching them with a dry pipette tip before reading
4. 4 Allow plate to stand 1-2 minutes after reagent addition for bubbles to dissipate

Related Video (3)

Bilibili (China-Accessible Mirrors) ★ 82

ELISA Operation Demonstration Video

"Concise hands-on ELISA benchwork demonstration showing proper operational technique, including the pipetting and plate handling steps where bubble formation typically occurs."

Bilibili (China-Accessible Mirrors) ★ 78

ELISA Experimental Protocol Complete Series (5 Episodes)

"Comprehensive 5-episode ELISA protocol series with detailed benchwork that would show detergent-containing wash/blocking steps and correct pipetting to avoid bubbles."

Bilibili (China-Accessible Mirrors) ★ 74

R&D Systems Quantikine ELISA Operation Guide

"Official Quantikine ELISA walkthrough covering sample preparation through detection, illustrating the detergent-containing buffers and plate-reading stages where bubbles cause high CV."