Improving Estimation of Intervention Impact on Antibody Titer Increase in the Presence of Missing Values: A Proposed Method for Addressing Limit of Detection Issues

In many laboratory assays that measure immunological quantities, a portion of the measured values fall below a limit of detection (LOD). This is also the case for the hemagglutination inhibition assay (HAI), a common method used to quantify antibodies in influenza research. The conventional approach is to treat values below the LOD as either equal to the LOD or LOD/2, which can introduce potential biases. These biases can become more pronounced when calculating compound measures such as the difference between post-vaccination and pre-vaccination antibody titers (titer increase). To address this issue, we conducted simulations using LOD measurements with LOD/2 values as the standard imputation. We then developed a new method to adjust coefficient estimates that account for the censored nature of measurements below the LOD. Applying this new method to data from an influenza vaccine cohort study, we compared the impact of vaccine dose on the titer increase of HAI. Author Summary Analysis of measurements obtained from widely used antibody assays frequently overlooks the underlying data structure, leading to potential biases in the results. To address this issue, we have developed a method that effectively reduces these biases. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement Yang Ge was supported by the Start-up Grant from the University of Southern Mississippi and NIH contract 75N93019C00052. Ted M. Ross is supported by the Georgia Research Alliance as an Eminent Scholar. Andreas Handel received partial support from NIH grants/contracts U01AI150747, R01AI170116, 75N93019C00052 and 75N93021C00018. Ye Shen received partial support from NIH grants/contracts R35GM146612, R01AI170116 and 75N93019C00052. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The study collected the data received the approval from the Western Institutional Review Board and the Institutional Review Boards of the University of Pittsburgh and the University of Georgia. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes All data of this study are provided in the Supplementary Materials.