From 2676115f77f0052902e1dcc0632420341464373d Mon Sep 17 00:00:00 2001
From: Mike Vink <mike1994vink@gmail.com>
Date: Tue, 27 Apr 2021 18:31:00 +0200
Subject: checkpoint 27-04-21

---
 data_understanding/main.tex | 580 ++++++++++++++++++++++++++++++++++----------
 1 file changed, 446 insertions(+), 134 deletions(-)

(limited to 'data_understanding/main.tex')

diff --git a/data_understanding/main.tex b/data_understanding/main.tex
index ff544c0..4f0b965 100644
--- a/data_understanding/main.tex
+++ b/data_understanding/main.tex
@@ -17,8 +17,6 @@
 
 \subsection{Technical description data collection}
 
-\subsubsection{MySQL database set up and data import}
-
 By following the guide on the
 \href{https://github.com/LogIN-/fluprint}{FluPrint Github Repository} the MySQL
 server was set up. In this work the FluPrint github was first added as a
@@ -59,79 +57,40 @@ The following subsections will list the information required from the data per
 data mining goals that are needed to answer the following business questions:
 
 \begin{itemize}
-        \item Which datasets in the FluPrint database are most interesting?
-        \item How do different clinical studies compare?
-        \item What are the differences in efficacy between vaccination types?
-        \item What is the effect of repeat vaccination on vaccine response?
-        \item What immunological factors correlate to a high vaccine response?
+        \item What kind of studies can be done using the FluPRINT database?
+        \item What immunological factors correlate to a high vaccine responses?
 \end{itemize}
 
 \subsubsection{Requirements per data mining goal}
 
 \begin{displayquote}
-"Explore and describe SQL queries and corresponding csv tables."
+"Explore and describe the database and corresponding tables."
 \end{displayquote}
 
 Falling under this data mining objective are the outputs and tasks related to
 data collection and description. These comprise a report on the initial
-collection of the data, selection of data, and description of general
-properties of the data. The data in this case is in a database format, thus
-here we describe the tables, keys, and attributes in the database, and also
-include descriptive statistics about the data. The goal is to replicate the
-description done in \cite{tomicFluPRINTDatasetMultidimensional2019} as well.
-Using these descriptions we provide insight into which datasets in the database
-are most interesting, and why in \cite{tomicSIMONAutomatedMachine2019} one
-dataset in particular was chosen.
-
-\begin{displayquote}
-"Model and visualise the different clinical study populations."
-\end{displayquote}
-\begin{displayquote}
-"Model and visualise the difference between vaccination types."
-\end{displayquote}
-\begin{displayquote}
-"Model and visualise repeat vaccination effects."
-\end{displayquote}
-
-In order to answer the business question "How do clinical studies compare?"
-subpopulations and groups of attributes need to be visualised and compared
-across different clinical studies. The data required must have rows
-corresponding to donors in a particular clinical study and columns that are
-attributes of tables in the database, these could be biological assay results
-or information about the donors. Thus we aimed to export one csv from the
-database per clinical study by querying for different clinical studies.
-
-We aimed to generate csv files of donors corresponding to received vaccine
-types to answer the business question "What are the differences in efficacy
-between vaccination types?". One simple method to indicate the difference
-between vaccines would be to report the proportion of high-reponders across all
-donors, or to use a simple model to find the best predictor for a high
-response. These comparisons require one table per vaccine type, with rows
-corresponding to donors and columns that include the vaccine response
-classification, in addition to other immune assay and donor attributes.
-
-The objective in question "What is the effect of repeat vaccination on vaccine
-response?" requires data from long running clinical studies. One dataset that
-is used by the database authors and was investigated to answer this question
-was already available, here we aimed to describe and visualise any patterns we
-could find in this dataset and other long running clinical study datasets. This
-required data from a subset of clinical studies that spanned multiple years, at
-this point in the project the data for these clinical studies should have been
-available, and we just had to choose those that spanned multiple years.
+collection of the data, selection of data, and description of properties of the
+data. The data in this case is in a database format, thus here we describe the
+tables, keys, and attributes in the database, and provide descriptive analyses
+where possible. The goal is to replicate the description done in
+\cite{tomicFluPRINTDatasetMultidimensional2019}, and to provide a more detailed
+explanation of the database from a user perspective.  Using these descriptions
+we provide insight into what kind of studies are possible with the database,
+and why the initial dataset in \cite{tomicSIMONAutomatedMachine2019} was chosen.
 
 \begin{displayquote}
-"Apply standard feature selection methods to the most interesting dataset."
+"Apply standard feature selection methods to the most interesting datasets."
 \end{displayquote}
 
 \begin{displayquote}
-"Fit classification models to the most interesting dataset."
+"Fit classification models to the most interesting datasets."
 \end{displayquote}
 
-These last two data mining objectives were chosen to comprise the data
-preparation and modelling phases of this project. The authors of fluprint set
-up an automated machine learning pipeline to investigate the longest running
-clinical dataset in the database. In this work we use a conventional data
-mining modelling process to replicate these results.
+These two data mining objectives were chosen to comprise the data preparation
+and modelling phases of this project. The authors of fluprint set up an
+automated machine learning pipeline to investigate the immunological factors
+that are correlated with a high vaccine response. In this work we use a
+conventional data mining modelling process to investigate these results.
 
 \section{Data description}
 
@@ -145,34 +104,37 @@ from 740 healthy donors, enrolled in influenza vaccine studies conducted by the
 Stanford-LPCH Vaccine Program from 2007 to 2015. These studies are described in
 the table accompanying the online publication of the fluprint dataset
 \autoref{tbl:studiesDesc}.  From those 740 donors a vaccine response
-classification was only given for 372 donors \autoref{fig:demoResponder}, by a
+classification was only given for 372 donors \autoref{fig:demoGraph}, by a
 method that will be described in the section describing the data table
 containing this attribute. Overall there was no major difference in demographic
-statistics when stratisfying the data in high or low responders
-\autoref{fig:demoResponder}.
-
-Importantly in all studies the donors are only vaccinated once, except in the
-study SLVP015, participants here were vaccinated annually from 2007-2015
-\autoref{tbl:studiesDesc}. In all other studies participants would only be
-studied within the scope of one influenza season.
-
-\fptable{studies_table}{.7}
-{Reference table of clinical studies}
-{Clinical study ID used (but remapped) in the database, age information,
-vaccine type information, and assay data types of clinical studies are in the
-rest of the columns.}
-{tbl:studiesDesc}
-
+statistics when stratisfying the data in high or low responder classification
+\autoref{fig:demoGraph}.
+
+Importantly, it is reported that in all studies the donors are only vaccinated
+once, except in the study SLVP015 \autoref{tbl:studiesDesc}
+\citep{tomicFluPRINTDatasetMultidimensional2019}. However, in later work of the
+same authors it is claimed that vaccines are administered as specified by the study
+\citep{tomicSIMONAutomatedMachine2019}.
+
+The donors for which a vaccine respone classification was available from all
+clinical studies together span a wide age range \autoref{fig:demoGraph}A from 1
+- 50 \autoref{tbl:demoStats}, in the original work the demographic statistics
+include the donors for which no vaccine response classification is given,
+therefore they report a greater range of 1-90. Stratisfying the donors on
+vaccine response does not affect the demographic attribute distribution, but
+the maximum age is lowered in the high responders group
+\autoref{fig:demoGraph}B.
 
 \begin{figure}
     \includegraphics[width=\textwidth]{demographic}
-    \label{fig:demoResponder}
     \caption{\textbf{A.} percentage of donors with factor property within high
     and low responder groups. Included are sex, race, and CMV status
     information. \textbf{B.} Age distribution of donors with a known response
-    classification.}
+    classification.}\label{fig:demoGraph}
 \end{figure}
 
+
+
 \begin{table}
 \centering
 \begin{tabular}{ll}
@@ -197,66 +159,416 @@ Other (\%) & 121  ( 32.5 )\\
 Unknown (\%) & 2  ( 0.5 )\\
 \bottomrule{}
 \end{tabular}
-    \caption{\textbf{Demographic statistics of donors with known vaccine response classification.}}
+\caption{\textbf{Demographic statistics of donors with known vaccine response classification.}}\label{tbl:demoStats}
 \end{table}
 
+\fptable{studies_table}{.7}
+{Reference table of clinical studies}
+{Clinical study ID used (but remapped) in the database, age information,
+vaccine type information, and assay data types of clinical studies are in the
+rest of the columns.}
+{tbl:studiesDesc}
+
+
+The data from the clinical studies consisted of 121 CSV files that were
+imported into the FluPrint database. The data was used to build four tables
+which will be described in the next sections, but we will not discuss technical
+validation of the database construction, refer to the original work for that
+\citep{tomicFluPRINTDatasetMultidimensional2019}.  The relation between the
+tables is best visualised in the original work of
+\citep{tomicFluPRINTDatasetMultidimensional2019}, it describes the MySql
+attribute types and columns in the tables \autoref{fig:tablesFluprint}
+(copied). The volume of the data is also given in the original work, per table
+the number of rows and columns is reported \autoref{tbl:volumeTables}.
+
+\begin{figure}
+    \includegraphics[width=\textwidth]{tablesFluprint}
+    \caption{
+        \textbf{(taken from original paper)} The FluPRINT database model. The diagram shows a schema of the FluPRINT
+    database. Core tables, donors (red), donor\_visits (yellow),
+    experimental\_data (blue) and medical\_history (green) are interconnected.
+    Tables experimental\_data and medical\_history are connected to the core
+    table donor\_visits. The data fields for each table are listed, including
+    the name and the type of the data. CHAR and VARCHAR, string data as
+    characters; INT, numeric data as integers; FLOAT, approximate numeric data
+    values; DECIMAL, exact numeric data values; DATETIME, temporal data values;
+    TINYINT, numeric data as integers (range 0–255); BOOLEAN, numeric data with
+    Boolean values (zero/one). Maximal number of characters allowed in the data
+    fields is denoted as number in parenthesis.
+    }\label{fig:tablesFluprint}
+\end{figure}
+
+\begin{table}
+    \centering
+    \begin{tabular}{lll}
+        \toprule{}
+        \textbf{Table name} & \textbf{Rows} & \textbf{Columns} \\
+        \midrule{}
+        \textit{donors} &  740 & 6 \\
+        \textit{donor\_visits} & 2,937 & 18 \\
+        \textit{experimental\_data} & 371,260 & 9 \\
+        \textit{Medical history} & 740 & 18 \\
+    \bottomrule{}
+    \end{tabular}
+    \caption{Volume of tables in the Fluprint database.}\label{tbl:volumeTables}
+\end{table}
+
+\subsection{Attribute types and values}
 
-\citep{chattopadhyaySinglecellTechnologiesMonitoring2014}
-The complex heterogeneity of cells, and their interconnectedness with each
-other, are major challenges to identifying clinically relevant measurements
-that reflect the state and capability of the immune system. Highly multiplexed,
-single-cell technologies may be critical for identifying correlates of disease
-or immunological interventions as well as for elucidating the underlying
-mechanisms of immunity. Here we review limitations of bulk measurements and
-explore advances in single-cell technologies that overcome these problems by
-expanding the depth and breadth of functional and phenotypic analysis in space
-and time. The geometric increases in complexity of data make formidable hurdles
-for exploring, analyzing and presenting results. We summarize recent approaches
-to making such computations tractable and discuss challenges for integrating
-heterogeneous data obtained using these single-cell technologies.
-
-\citep{galliEndOmicsHigh2019}
-High-dimensional single-cell (HDcyto) technologies, such as mass cytometry
-(CyTOF) and flow cytometry, are the key techniques that hold a great promise
-for deciphering complex biological processes. During the last decade, we
-witnessed an exponential increase of novel HDcyto technologies that are able to
-deliver an in-depth profiling in different settings, such as various autoimmune
-diseases and cancer. The concurrent advance of custom data-mining algorithms
-has provided a rich substrate for the development of novel tools in
-translational medicine research. HDcyto technologies have been successfully
-used to investigate cellular cues driving pathophysiological conditions, and to
-identify disease-specific signatures that may serve as diagnostic biomarkers or
-therapeutic targets. These technologies now also offer the possibility to
-describe a complete cellular environment, providing unanticipated insights into
-human biology. In this review, we present an update on the current cutting-edge
-HDcyto technologies and their applications, which are going to be fundamental
-in providing further insights into human immunology and pathophysiology of
-various diseases. Importantly, we further provide an overview of the main
-algorithms currently available for data mining, together with the conceptual
-workflow for high-dimensional cytometric data handling and analysis. Overall,
-this review aims to be a handy overview for immunologists on how to design,
-develop and read HDcyto data.
-
-\citep{simoniMassCytometryPowerful2018}
-Advancement in methodologies for single cell analysis has historically been a
-major driver of progress in immunology. Currently, high dimensional flow
-cytometry, mass cytometry and various forms of single cell sequencing-based
-analysis methods are being widely adopted to expose the staggering
-heterogeneity of immune cells in many contexts. Here, we focus on mass
-cytometry, a form of flow cytometry that allows for simultaneous interrogation
-of more than 40 different marker molecules, including cytokines and
-transcription factors, without the need for spectral compensation. We argue
-that mass cytometry occupies an important niche within the landscape of
-single-cell analysis platforms that enables the efficient and in-depth study of
-diverse immune cell subsets with an ability to zoom-in on myeloid and lymphoid
-compartments in various tissues in health and disease. We further discuss the
-unique features of mass cytometry that are favorable for combining multiplex
-peptide-MHC multimer technology and phenotypic characterization of antigen
-specific T cells. By referring to recent studies revealing the complexities of
-tumor immune infiltrates, we highlight the particular importance of this
-technology for studying cancer in the context of cancer immunotherapy. Finally,
-we provide thoughts on current technical limitations and how we imagine these
-being overcome.
+Because of the great number of attributes in the database, we discuss them by
+table starting with the donors \autoref{fig:tablesFluprint}.
+
+\subsubsection{donors table}
+
+The \textit{donors.id} attribute is simply an enumeration of unique donors,
+importantly, it is used as a key to get attributes from other tables. The
+column \textit{study\_donor\_id} is an encrypted identification number. Each
+donor belongs to the study identified by the \textit{study\_id}, these are the
+last two digist of the name code (those starting with SLVP0 \(\cdot\cdot\)) in
+the reference table \autoref{tbl:studiesDesc}, the \textit{study\_internal\_id}
+is either the digit or a string containing the digit in \textit{study\_id}. The
+\textit{gender} and \textit{race} attribute contain the values used in
+\autoref{fig:demoGraph}, a minor note is that in the original paper "American
+Indian or Alaska Native" is listed as one of the \textit{race} values but is
+not used in the database. There are 5 donors whose race is "NULL", which are
+mapped to unkown \autoref{fig:demoGraph}.
+
+\begin{table}
+    \begin{tabular}{rlrlll}
+\toprule{}
+id & study\_donor\_id & study\_id & study\_internal\_id & gender & race\\
+\midrule{}
+1 & e27ad74ff9a5f2f32d8e852533f054c0 & 30 & 30 & Female & Asian\\
+2 & 4a89ac4d3f4dc869e5c8e8cf862cffda & 30 & 30 & Male & Other\\
+3 & a2cde6e54dec92422b0427dd49244350 & 30 & 30 & Female & Caucasian\\
+4 & 0f7d8d1c13e876017ea465f99d25581f & 30 & 30 & Male & Other\\
+5 & 1ed2f6409584b7b4e9720b28d794fe91 & 30 & 30 & Female & Caucasian\\
+\addlinespace
+6 & a575678405e9615bfb87eccfa031f7fc & 30 & 30 & Male & Other\\
+\bottomrule{}
+\end{tabular}
+    \caption{Head of the donors table.}\label{tbl:donorsHead}
+\end{table}
+
+\subsubsection{donor\_visits table}
+
+The donor visits table is the core table of the database, it contains donor
+attributes at visit times during enrolment in clinical studies in rows that are
+uniquely identified by an \textit{id} integer. Each
+row also includes the \textit{donor\_id} identify the donor that visitted.
+
+The database combines different clinical studies accross years and the data
+from these studies is incomplete leading to an incomplete and hetergenous
+database \autoref{tbl:visitsDesc}. For example some donors might miss their
+second visit to determine their antibody levels, or the number of parameters
+measured by an assay changed in the timespan of a clinical study. Unifying
+these clinical studies in one database resulted in normalised but incomplete
+data and heterogenous data. More specifically, every attribute in the core
+table has missing value, which complicates dataset selection. One examples of
+visit data of a donor is discussed to highlight important attributes and
+problems in the data: that the number of visits is variable, that all columns
+are incomplete, and that classification is sometimes based on single visits or
+inconsistent \autoref{tbl:visit166} \autoref{tbl:visitsDesc}.
+
+\begin{table}
+\addtolength{\leftskip} {-2cm} % increase (absolute) value if needed
+\addtolength{\rightskip} {-2cm} % increase (absolute) value if needed
+\begin{tabular}{lrrrrrrrrr}
+\toprule{}
+stat & age & cmv\_status & ebv\_status & bmi & vaccine & geo\_mean & d\_geo\_mean & vaccine\_resp & total\_data\\
+\midrule{}
+n & 2937.0 & 1081.0 & 548.0 & 516.0 & 2794.0 & 984.0 & 1260.0 & 1206.0 & 2937.0\\
+na & 0.0 & 1856.0 & 2389.0 & 2421.0 & 143.0 & 1953.0 & 1677.0 & 1731.0 & 0.0\\
+mean & 47.3 & 0.4 & 0.8 & 24.8 & 3.7 & 87.6 & 8.9 & 0.3 & 126.4\\
+sd & 27.0 & 0.5 & 0.4 & 5.6 & 1.0 & 101.7 & 30.9 & 0.4 & 368.4\\
+se\_mean & 0.5 & 0.0 & 0.0 & 0.2 & 0.0 & 3.2 & 0.9 & 0.0 & 6.8\\
+\addlinespace
+IQR & 50.2 & 1.0 & 0.0 & 6.7 & 0.0 & 105.4 & 4.0 & 1.0 & 19.0\\
+skewness & 0.2 & 0.3 & -1.4 & 1.0 & -1.7 & 3.6 & 9.9 & 1.1 & 7.1\\
+kurtosis & -1.5 & -1.9 & -0.1 & 2.1 & 3.0 & 26.6 & 114.9 & -0.9 & 49.7\\
+\bottomrule{}
+\end{tabular}
+\caption{Descriptive stats of relevant numeric or binary factor columns in the
+    donor visits table. For geo\_mean 0 is considered as missing data.}\label{tbl:visitsDesc}
+\end{table}
+
+Per donor all visits are enumerated in chronological order by
+\textit{visit\_id} \autoref{tbl:visit166}. Further visit info includes:
+\textit{visit\_internal\_id} which is a number that indicates the visit order
+within an influenza season but this differs per clinical study (e.g. some use
+1-2-3, orther use 0-7-28), the \textit{vist\_year} is the influenza season of
+the visit, the \textit{visit\_day} is the number of days relative to the date
+of vaccination,  \textit{age} and \textit{age\_round} indicate the donor's age
+at time of the visit, and \textit{bmi} gives the donor bmi at visit time, and
+lastly \textit{visit\_type\_hai} is the intent of the visit which is either
+"pre", "post", or "other",
+
+During the "pre" visit a virological assay is performed to determine the CMV
+and Epstein-Barr virus (EBV) status of the donor, which are indicated by the
+binary variables \textit{cmv\_status} and \textit{ebv\_status}.
+
+To measure vaccine response to a vaccine which is indicated by an id
+\autoref{tbl:remapVaccine} in \textit{vaccine}, the hemagglutination inhibition
+assay (HAI assay) is used. The procedure measures the influenza antibody titers
+before vaccination during the \textit{visit\_type\_hai} "pre" visit of a
+participant, and 28 days after vaccination during a "post" visit. The geometric
+mean titer (GMT) at each visit is calculated, and a fold change in GMT is
+calculated as the ratio of the GMT at day 28 (post) and during the first visit
+(pre). These values are \textit{geo\_mean} and \textit{d\_geo\_mean},
+\textit{d\_single} is the antibody titer fold-change per strain of virus used
+in the vaccine, it is unclear how this value is aggregated over different
+strains and is left out of further analysis.  This data was used to classify
+donors in high or low responders according to FDA guidelines \cite{},
+individuals are high-responders if they seroconverted (4-fold or greater rise
+in HAI titer) and were seroprotected (GMT HAI \(\ge\) 40) after vaccination.
+The seasonal vaccine response classifications are given by the binary variable
+\textit{vaccine\_resp}.
+
+The assays performed to get a serological/immunlogical profile of the donor
+before vaccination are described later in the section of the experimental data
+table, all assays are listed in the original work
+\cite{tomicFluPRINTDatasetMultidimensional2019} and are summarised here
+\autoref{tbl:assays}, the total rows of assay data is given by
+\textit{total\_data}.
+
+\begin{table}
+\addtolength{\leftskip} {-2cm} % increase (absolute) value if needed
+\addtolength{\rightskip} {-2cm} % increase (absolute) value if needed
+\begin{tabular}{rrrlrrrlrrrrr}
+\toprule{}
+visit\_id & year & day & type & age & cmv & ebv & bmi & vaccine & geo\_mean & d\_geo\_mean & response & assay\_data\_rows\\
+\midrule{}
+1 & 2011 & 0 & pre & 20 & 1 & 1 & 30.31 & 4 & 25.20 & 6 & 0 & 343\\
+2 & 2011 & 7 & other & 20 & 1 & 1 & NULL & 4 & 0.00 & 6 & 0 & 51\\
+3 & 2011 & 28 & post & 20 & 1 & 1 & NULL & 4 & 160.00 & 6 & 0 & 51\\
+4 & 2012 & 0 & pre & 21 & 1 & 1 & 30.31 & 4 & 9.28 & 4 & 0 & 292\\
+6 & 2013 & 0 & pre & 22 & 1 & 1 & 30.31 & 4 & 15.91 & 2 & 0 & 2877\\
+\addlinespace
+7 & 2013 & 7 & other & 22 & 1 & 1 & NULL & 4 & 0.00 & 2 & 0 & 63\\
+8 & 2013 & 28 & post & 22 & 1 & 1 & NULL & 4 & 26.75 & 2 & 0 & 82\\
+\bottomrule{}
+\end{tabular}
+\caption{Visit data of donor 166 from study SLVP021 \autoref{tbl:studiesDesc},
+where participants are only vaccinated once.
+Number of visits and data collected at visit varies, classification is
+inconsistent with \( \geq 40\) and 4-fold increase
+rule in 2011.}\label{tbl:visit166}
+\end{table}
+
+The most important data related to the visits of donor 166 is shown in Table
+\ref{tbl:visit166}.  The vaccine response classification is calculated based on
+the GMT in the "pre" and "post" visits.  This classification is done per
+influenza season, but the HAI assay requires a "pre" visit and a "post" visit
+28 days later to measure the difference in GMT.  However, sometimes a
+classification is given when there is only one visit record in a season, like
+in 2012 for donor 166 \autoref{tbl:visit166}.
+
+\begin{figure}
+    \includegraphics[width=\textwidth]{season_classification}
+    \caption{}\label{fig:classInconsistent}
+\end{figure}
+
+The example of donor 166 contains an inconsistency in the classification, in
+2011 the GMT \textit{geo\_mean} increases from 25.20 to 160.00, and the
+\textit{d\_geo\_mean} is 6, but in this season the donor is wrongly classified
+as a low responder \autoref{tbl:visit166}. Because of this the seasonal
+classification of donors was investigated using the seroprotection and
+seroconversion criteria \ref{fig:seasonalClasses}, records of incorrectly
+labelled donors are also saved as a spreadsheet.
+
+\subsubsection{Experimental data table}
+
+\begin{table}
+    \begin{tabularx}{\textwidth}{Xp{0.5\textwidth}X}
+\toprule{}
+        \textbf{Name} & \textbf{Description} & \textbf{id} (\textit{experimental\_data.assay})\\
+\midrule{}
+        (Multiplex) cytokine assays & Multiplex ELISA using Luminex polysterene
+        bead or magnetic bead kits. Measures serum cytokine/hormone level in
+        z.log2 units using fluorescent antibodies. & 3, 6, 15, 16\\
+        \addlinespace
+        Flow and mass cytometry assays & uses labeled antibodies to detect antigens on
+        a cell surface to identify a subset of a cell population, units are in
+        percentage of parent population. & 4, 9, 13, 17 \\
+        \addlinespace
+        Phosphorylation cytometry assays & Uses antibodies to measure
+        phosphorylation of specific proteins stimulated by an immune system event
+        belonging to cell population subsets. Units are a fold change between
+        stimulated and un-stimulated cells, for mass cytometry arcsin readout difference,
+        fold-change of 90th percentile readout values otherwise.  & 7, 10 (mass cytometry) (flow cytometry)\\
+        \addlinespace
+        complete blood count (CBCD) & Different cells are counted using flow
+        cytometry Units are usually in Count/$\mu$L & 11 \\
+        \addlinespace
+        meso scale discovery assays (MSD) & A setup where serum cytokines or hormones
+        are captured with antibodies, and then detected by using a detection
+        antibody. Units are arbitrary intensity & 2, 12, 14 \\
+\bottomrule{}
+\end{tabularx}
+    \caption{assays table}\label{tbl:assays}
+\end{table}
+
+
+Assays performed in visits are remapped, but the values in the
+database do not correspond to the reported table \autoref{tbl:remapVaccine}.
+Actual assay type, data units, and id in the database are reported here
+\autoref{tbl:assays}.
+
+\fpfig{exp_data_numbers}{.7}
+{Feature count per individual assay id, assay type, stratisfied in either response status or study}
+{caption}
+{fig:featureNumbers}
+
+In total there are data from 14 different assays, not counting the virological
+and HAI antibody assays \autoref{tbl:assays}. The virological assays include
+the cmv virus status and ebv status, and is not used in this work because it is
+done in a smaller subset of studies. Those 14 assays have been aggregated in
+this work to 5 different types of experiments: the multiplex assays measure
+serum molecules such as cytokines and other signaling molecules, flow and mass
+cell cytometry measure the phenotype of specific immune related cells,
+phosphorylation flow and mass cytometry measures the phosphorylation signaling
+pathway activation after an immune stimulation, the blood count measures the
+count of cells in the blood, and meso scale discovery (MSD) measures hormones
+or cytokines from the blood.
+
+\begin{figure}
+    \includegraphics[width=\textwidth]{assay_value_distributions}
+    \caption{noise in 90th \%tile}\label{fig:assayDistr}
+\end{figure}
+
+The experimental data table contains all features recorded for a donor visit.
+The number of features collected for each visit is large and varies greatly
+(mean at 126 , \(\pm \)368 SD) \autoref{tbl:visitsDesc}, and in total there are
+3285 different features measured across all clinical studies. However, not
+every assay is done in every clinical study \autoref{fig:featureNumbers} and
+over the years the data generated by assays has changed, so a table with all
+features as columns and all donors as rows would be extremely sparse (and
+crashes R due to RAM limitations).  Describing the 3285 different
+features in this sparse table would be impossible, but assay value
+distributions across studies are shown to follow normal or power distributions
+\autoref{fig:assayDistr}. Global correlation analysis is complicated by the
+great number of features and sparseness in the data.
+
+\begin{figure}
+    \includegraphics[width=\textwidth]{repeat_visits_per_study}
+    \caption{the number of donors that visited per number of influenza seasons
+    they visited (years), per study. The color indicates the number of visits for which a
+    classification was available, counted within the groups of donors that
+    visited the same amount of times.}\label{fig:repeatVisits}
+\end{figure}
+
+What further complicates selecting data is repeat visits of donors, and missing
+visits. The problem of repeat visits over a span of multiple influenza seasons
+is that not the same assay types are done, and that repeat visits are only a
+small portion of the database. The data is also not suitable right away for
+studying the effect of repeat vaccination on high versus low vaccine reponse,
+since the classification in the longitudal study (SLVP015) is mostly not
+available \autoref{fig:repeatVisits}.
+
+For example exploring the effect repeat vaccination has an response rate would
+first require manual labelling of high and low responses, at least for the
+cases where it is possible based on the GMT data. Those cases are when
+classification is set to a null value even though GMT data is available.  The
+reason for this null value assignment is reported, but the pattern seems to set
+the vaccine response to null if there is not enough assay data measured.
+
+\section{Data quality}
+
+The database has issues that are inherent to combining multiple studies and the
+classification is inconsistent in some cases \autoref{fig:classInconsistent},
+or often missing completely because no HAI antibody assay data was available or
+the classification was set to a null value by the database authors
+\autoref{fig:repeatVisits}.  The main value of the database is the assay data
+that is fully represented in all studies and across all years, but this
+information is hard to access since all studies do not use overlapping assays
+\autoref{fig:featureNumbers}, resulting in high sparsity data. Further, the
+sample size that can be used for further studies is limitted, since the high
+versus low vaccine response is only available for a small subset of the data.
+
+Specific attributes that have great amounts of missing values are the
+virological and HAI assay data, the last is used for the vaccine response
+classifcation. Potential for Studying the correlation of these values with
+vaccine response is thus limitted. Nevertheless assay data is often available
+and could be used to identify immunological factors that correlate with other
+data, such as repeat vaccination, the exploration of this effect is outside the
+scope of this work due to the data sparsity issues.
 
 \printbibliography
+
+\begin{appendices}
+    \section{Remaps used in the database}
+
+    \begin{table}[h]
+        \begin{tabular}{lll}
+            \toprule{}
+            Vaccine received & Vaccine type ID & Vaccine type name \\
+            \midrule{}
+            FluMist IIV4 0.2 mL intranasal spray & 1 & Flumist \\
+            FluMist Intranasal spray & 1 & Flumist \\
+            FluMist Intranasal Spray 2009–2010 & 1 & Flumist \\
+            FluMist Intranasal Spray & 1 & Flumist \\
+            Flumist & 1 & Flumist \\
+            Fluzone Intradermal-IIV3 & 2 & Fluzone Intradermal \\
+            Fluzone Intradermal & 2 & Fluzone Intradermal \\
+            GSK Fluarix IIV3 single-dose syringe & 3 & Fluarix \\
+            Fluzone 0.5 mL IIV4 SD syringe & 4 & Fluzone \\
+            Fluzone 0.25 mL IIV4 SD syringe & 5 & Paediatric Fluzone \\
+            Fluzone IIV3 multi-dose vial & 4 & Fluzone \\
+            Fluzone single-dose syringe & 4 & Fluzone \\
+            Fluzone multi-dose vial & 4 & Fluzone \\
+            Fluzone single-dose syringe 2009–2010 & 4 & Fluzone \\
+            Fluzone high-dose syringe & 6 & High Dose Fluzone \\
+            Fluzone 0.5 mL single-dose syringe & 4 & Fluzone \\
+            Fluzone 0.25 mL single-dose syringe & 5 & Paediatric Fluzone \\
+            Fluzone IIV3 High-Dose SDS & 6 & High Dose Fluzone \\
+            Fluzone IIV4 single-dose syringe & 4 & Fluzone \\
+            Fluzone High-Dose syringe & 6 & High Dose Fluzone \\
+            \bottomrule{}
+        \end{tabular}
+        \caption{Remaps of vaccine type relevant to to the clinical studies
+        reference table \autoref{tbl:studiesDesc}, and the section on the donor
+        visits table.}\label{tbl:remapVaccine}
+    \end{table}
+
+    \begin{table}
+        \begin{tabular}{ll}
+            \toprule{}
+            Original & Remapped \\
+            \midrule{}
+            No& 0 \\
+            Yes& 1 \\
+            IIV injection/im& 2 \\
+            Doesn’t know/doesn’t remember/na/does not remember& 3 \\
+            LAIV4 intranasal/laiv\_std\_intranasal/laiv\_std\_ intranasal/nasal/intranasal& 4 \\
+            \bottomrule{}
+        \end{tabular}
+        \caption{caption}\label{tbl:remapHistory}
+    \end{table}
+
+    \begin{table}
+        \begin{tabular}{ll}
+            \toprule{}
+            Original & Remapped \\
+            \midrule{}
+            CMV EBV & 1 \\
+            Other immunoassay & 2 \\
+            Human Luminex 62–63 plex & 3 \\
+            CyTOF phenotyping & 4 \\
+            HAI & 5 \\
+            Human Luminex 51 plex & 6 \\
+            Phospho-flow cytokine stim (PBMC) & 7 \\
+            pCyTOF (whole blood) pheno & 9 \\
+            pCyTOF (whole blood) phospho & 10 \\
+            CBCD & 11 \\
+            Human MSD 4 plex & 12 \\
+            Lyoplate 1 & 13 \\
+            Human MSD 9 plex & 14 \\
+            Human Luminex 50 plex & 15 \\
+            Other Luminex & 16 \\
+            \bottomrule{}
+        \end{tabular}
+        \caption{caption}\label{tbl:remapAssays}
+    \end{table}
+\end{appendices}
+
 \end{document}
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