Multiplexed Nucleic Acid Programmable Protein Arrays

Yu, Xiaobo; Song, Lusheng; Petritis, Brianne; Bian, Xiaofang; Wang, Haoyu; Viloria, Jennifer; Park, Jin; Bui, Hoang; Li, Han; Wang, Jie; Liu, Lei; Yang, Liuhui; Duan, Hu; McMurray, David N.; Achkar, Jacqueline M.; Magee, Mitch; Qiu, Ji; LaBaer, Joshua

Finally, we evaluated the reproducibility of M-NAPPA arrays for protein array preparation and protein-protein interactions. We found M-NAPPA can be reproducibly fabricated with spot-to-spot, zone-to-zone and slide-to-slide CVs that are similar to those obtained with NAPPA ( Table S4) . The spot-to-spot and zone-to-to zone CVs for protein-protein interactions were also similar between the two array platforms (Table S5) . While the correlations within and between different M-NAPPA slides were good (i.e., R = 0.93 for both) (Figure 3 and Figure S6 ), with some size adaptation, the reproducibility could eventually be further improved with the use of automation equipment like the HS 4800 Pro Hybridization Station (Tecan Trading AG; Männedorf, Switzerland). In some ways, M-NAPPA resembles "natural protein" microarrays that print unpurified or partially fractionated proteins from lysates of human cells, tissues or body fluids, but in a much more controlled manner. Each feature of a natural protein microarray typically represents a mixture of unknown proteins. Thus, responsive hits on natural protein arrays require a challenging and time-consuming process to determine the identity of the protein responsible for the response. This may require further purification, identification by mass spectrometry and additional response testing of recombinant proteins [46, 47] . In the case of M-NAPPA, the identities of the proteins in each mix are known in advance and the plasmids encoding for each protein are available for secondary testing.