HAI Antigen Unit Standardization
Prepare a caller-specified HAI antigen-unit input by volume dilution of a measured HA stock; assay-specific validation remains required.
Description
Calculate stock and diluent volumes for a target hemagglutinating-unit amount within a specified final volume.
Hemagglutinating units, abbreviated HAU, describe an assay-defined agglutinating activity. An HAI assay uses antigen standardized to a defined activity amount under its protocol. HAU are not automatically PFU, particle counts, or antibody titers.1
A stock value expressed as HAU per reference volume can be converted into activity per microliter. The desired unit amount then determines the volume of stock needed; the remaining final volume is diluent. The reference volume matters as much as the numerical unit count.
HAI Antigen Unit Standardization returns stock volume, diluent volume, and a dilution factor. It performs a dilution calculation and rejects a target that would require more stock than the specified final volume.
When to check an antigen dilution calculation
Use the calculator after stock HAU have been measured under the applicable assay conditions and target units chosen. It helps keep the reference-volume convention explicit during a concentration-to-volume calculation.
- Calculate stock volume from HAU measured per reference volume.
- Check whether a target can be reached by dilution alone.
- Distinguish fold dilution from the stock fraction of final volume.
How to enter activity and calculate dilution volumes
- Enter positive Stock hau per reference volume and the corresponding positive Reference volume ul.
- Enter positive Target hau and Target reaction volume ul for the final mixture being calculated.
- Run the tool and read Stock volume ul, Diluent volume ul, and Dilution factor.
- Keep the assay’s reference-volume basis and subsequent mixing steps with the calculation.
stockVolumeUl = targetHau×referenceVolumeUl/stockHauPerReferenceVolume; diluentVolumeUl = targetReactionVolumeUl − stockVolumeUl; dilutionFactor = targetReactionVolumeUl/stockVolumeUl The implementation assumes activity scales with volume under the chosen assay convention. The dilution factor is final volume divided by stock volume, not its reciprocal.
{
"input": {
"stockHauPerReferenceVolume": 64,
"referenceVolumeUl": 25,
"targetHau": 4,
"targetReactionVolumeUl": 25
},
"output": {
"stockVolumeUl": 1.5625,
"diluentVolumeUl": 23.4375,
"dilutionFactor": 16
}
}If stock volume equals final volume, diluent is zero and dilution factor is 1. A larger required stock volume is rejected rather than returning negative diluent. Nonpositive fields and nonfinite inputs produce errors.
Limitations and interpretation
- The calculator does not measure HAU, choose the assay target, validate antigen stability, or confirm a back-titration.
- Assay conditions and the reference-volume definition determine the meaning of the stock activity.
- No allowance is made for pipetting feasibility, dead volume, replicate scaling, or additional specimen/reagent volumes unless they are included in the supplied final-volume definition.
- Dilution alone cannot concentrate a stock below the required activity concentration.
Alternative or Complementary methods
Keep the measured HA endpoint with the stock activity record. A paired HAI titer comparison summarizes antibody inhibition results, which are separate from antigen unit standardization.
References
-
Manual for the laboratory diagnosis and virological surveillance of influenza — World Health Organization (2011)
Similar or alternative tools
- Hemagglutination Dilution Endpoint
Read a scored agglutination endpoint while retaining above-range censoring.
- Paired HAI Titer Rise
Calculate a ratio of acute and convalescent reciprocal antibody-inhibition titers.