Fig. 5

PSA detection from blood sample. a Flow chart illustrates steps for detecting PSA in blood sample: (1) Plasma platelets were separated from blood with centrifugation. (2) The plasma platelet layer was extracted and mixed with (anti-PSA)-MNPs. The (anti-PSA)-MNPs were magnetically separated and washed with PBS. (3) The (anti-PSA)-MNPs were added into cis-chamber of a flow cell. A potential difference was applied between both sides of the nanopore and a magnetic field was applied from trans-side of flow cell to initialise detection process. (4) Ionic current measured by patch clamp amplifier was analysed by an event classifier where three distinct event types—translocation, blocking and unblocking are detected and classified as they occur. This enables tracking of the number of blocking and unblocking events that occurred over the course of analysis. For application here in quantitative analysis, SiN membranes have been thinned down to about 80 nm. b Ionic current trace from one experiment that employed (anti-PSA)-MNPs to capture PSA from whole blood and transferred to the 3 × 3 array nanopore blockade sensor. The trace shows four blockade events but upon switching on the cis-magnet it transpires two of these blockade events are non-specific. c Irreversible blockades observed versus PSA concentrations. Blockade events were counted after 4 cycles of switching on trans-magnet for 10 min, and subsequently cis-magnet for 5 min to maximise probability of blockade events at extremely low concentrations. Mean value of irreversible blockades and 95% exact confidence intervals were obtained from a total of 31 measurements (n = 5, 10, 6, 5 and 5 from lower to higher concentrations; one measurement per chip). Of these measurements, 23 chips were employed and 8 of them were regenerated. The exact method was used to compute 95% confidence intervals for Poisson means. The difference between any two determined Poisson means was tested by a two-tailed z-test of square root transform version; ns = not significant, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001 and ****p ≤ 0.0001. d Comparison of PSA level from whole blood using nanopore blockade sensor versus an ELISA kit. Error bars represent uncertainty of determined PSA concentration at 95% confidence by nanopore sensing (n = 6) and ELISA kit (n = 12), respectively