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ELISA (Competitive) severe

B/B0 Ratio at Curve Endpoint Outside 5-95% Range

Symptom
The B/B0 ratio (bound/maximum bound signal) for the highest or lowest standard concentration exceeds 95% or falls below 5%, indicating the standard curve dynamic range does not adequately cover the analyte concentration range.
Common Causes
  1. 1 Standard curve range not optimally positioned for the competitive assay format
  2. 2 Highest standard concentration too low (B/B0 >95%)
  3. 3 Highest standard concentration too high (B/B0 <5%)
  4. 4 Improper standard dilution series preparation
Solutions
  1. 1 Increase the standard concentration by one dilution factor if B/B0 >95% at curve endpoint
  2. 2 Decrease the standard concentration by one dilution factor if B/B0 <5% at curve endpoint
  3. 3 Verify proper pipetting and serial dilution technique for standard preparation
  4. 4 Adjust the concentration range to position curve endpoints between 5-95% B/B0
Related Video (3)
Bilibili (China-Accessible Mirrors) ★ 80
How to Run an R&D Systems Quantikine ELISA
"Quantikine ELISA workflow video explicitly includes troubleshooting guidance relevant to standard curve and dynamic range issues."
Thermo Fisher Scientific ★ 72
How to Run an ELISA Assay – Invitrogen Kit Step-by-Step Tutorial
"Step-by-step kit tutorial demonstrates proper setup and standard curve generation essential for avoiding range positioning failures."
Bilibili (China-Accessible Mirrors) ★ 68
Thermo Fisher Microplate Reader Software Operation
"Microplate reader software operation covers data acquisition where B/B0 ratios and standard curve range are evaluated."
Source: abcam.com ↗
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