Calculate antibody titers from serial dilution data and determine optimal dilution ratios for immunoassays and western blotting applications.
Antibody Volume Needed
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Buffer Volume Needed
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Dilution Ratio
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Calculated Titer
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An antibody titer is the highest dilution of an antibody solution that still produces a detectable signal. It helps determine the optimal concentration for experiments and ensures reproducible results while minimizing background noise.
Start with the manufacturer's recommended dilution range, typically 1:100 to 1:1000 for primary antibodies. If no recommendation exists, begin with 1:100 and perform a serial dilution to find the optimal concentration.
Primary antibodies bind directly to your target protein and are typically used at 1:100-1:5000 dilutions. Secondary antibodies bind to primary antibodies and are usually more concentrated, requiring higher dilutions of 1:1000-1:10000.
Include 6-8 dilution points spanning 2-3 log orders to capture the full dynamic range. This ensures you find both the optimal working dilution and the point where signal drops to background levels.
Use 2-fold or 3-fold dilutions for fine titration when working near the optimal range. Use 5-fold or 10-fold dilutions for broader screening or when you need to cover a wider concentration range quickly.
Avoid storing diluted antibodies when possible, as dilution reduces stability. If storage is necessary, add BSA or glycerol as stabilizers, store at 4°C, and use within 24-48 hours for best results.
Use the formula C1×V1 = C2×V2, where C1 is stock concentration, V1 is stock volume needed, C2 is final concentration, and V2 is final volume. This calculator automates this calculation for you.