## 2 Materials and Methods
### SPRI Analysis  
#### 2.3.1 SPR Measurement  
The antibody-coated sensor surface was continuously illuminated by a fixed laser diode at a wavelength of 840 nm. The light passed through the prism, reflected on the back side of the sensor chip surface, and was captured by a CCD camera, which monitored the intensity of the reflected light over time for the regions of interest (ROIs); see Figure 1 for SPRi set-up. The images of the camera were visualized in real time using DAX software version 1 (IBIS technologies, Enschede, The Netherlands). Before analysis, the sensor was treated with two blocking buffers composed of 100 mmol/L 2-amino ethanol and 1%  
\begin{table}
\begin{tabular}{l l l l l} \hline \hline  & **Antibody** & **Isotype** &
\begin{tabular}{l} **Commercial** \\ **Concentration** \(\upmu\)g/mL** \\ \end{tabular} & **Manufacture** \\ \hline \multirow{6}{*}{**Oxidation biomarkers**} & Anti-Malondialdehydedehyde & Mouse monoclonal IgG1 & 1000 & Abcam, Cambridge, UK \\  & Anti-Malondialdehydedehyde & Rabbit polyclonal IgG & 1000 & Cloud-Clone Corp., Wuhan, \\  & Anti-4- & & & China \\  & hydroxynonenal & Rabbit polyclonal IgG & 500 & Cel biolabs Inc., San Diego, \\  & Anti-4- & & & CA, USA \\  & hydroxynonenal & Goat polyclonal IgG & 1000 & Cel biolabs Inc., San Diego, \\  & Anti-ditryrosine & Mouse monoclonal & & Japan institute for the \\  & Anti-dytrosine & IgG2a/k & 200 & controls of aging, Fukuroi, \\  & Anti-Advanced & & & Shizuoka, Japan \\  & glycation end product & Mouse monoclonal IgG1/k & 1000 & Cloud-Clone Corp., Wuhan, \\ \hline \multirow{6}{*}{**Body fluid-specific biomarkers**} & Anti-semenologin-1 & Mouse monoclonal IgG1 & 200 & Santa Cruz, CA, USA \\  & Anti-human-prostate- & Mouse monoclonal IgG1 & 1000 & Bio-Rad, Hercules, CA, USA \\  & specific-antigen & & & CA, USA \\  & Anti-human & & & CA, USA \\  & Tamm–Horsfall ascites & Mouse monoclonal IgG2b & 200 & Cedarlane, Burlington, ON, \\  & Anti-osteepontin & Mouse monoclonal IgG1/k & 1000 & Bivosion, Milpitas, CA, USA \\  & Anti-alpha & Mouse monoclonal & 400 & Abgent, San Diego, CA, USA \\  & amylase 1 & IgG2a/k & & CA, USA \\ \hline \multirow{6}{*}{**Isotype controls**} & IgG Isotype control & Rabbit polyclonal IgG & 1000 & Merck KGaA, Darmstadt, \\  & IgG1 Isotype control & Mouse monoclonal IgG1 & 500 & Germany \\  & & & & R\&D systems, Minneapolis, \\ \cline{1-1}  & IgG2a Isotype control & Mouse monoclonal IgG2a & 500 & MN, USA \\ \cline{1-1}  & IgG2b Isotype control & Mouse monoclonal IgG2b & 200 & Thermo Fisher Scientific, \\ \hline \hline \end{tabular}
\end{table}
Table 2: Overview showing the selected antibodies and respective isotypes that were used.  
(\(v/v\)) bovine serum albumin (BSA) (both Sigma-Aldrich, Darmstadt, Germany) to prevent non-specific binding to the sensor. The SPR measurement started with four regeneration runs, to wash and stabilize the sensor, followed by a sample run and ended with a single regeneration run before the injection of the next sample. Each cycle of the regeneration run consisted of three phases: (i) baseline phase of two minutes, (ii) association phase of 20 min with 120 \(\upmu\)L regeneration buffer (RB) (0.1M Glycine, pH \(\pm\) 3.0 (Merck, Darmstadt, Germany) and 0.05% Tween-20 (\(v/v\)) (Sigma-Aldrich, Darmstadt, Germany)), and (iii) dissociation phase of 8 min to wash the sensor with 240 \(\upmu\)L PBS. For the sample run, the same cycle was repeated, but during the association step, 150 \(\upmu\)L body fluid was injected instead of regeneration buffer. The sample was flowed back and forth over the sensor surface, enabling specific binding of the biomarkers in the sample to the immobilized antibodies. Subsequently, a single regeneration run was performed to remove the bound antigens to reverse the interaction between the antigens and immobilized antibodies and to return the response signals to baseline.