Paper based analytical devices (PADs) have great potential in the application of point-of-care diagnosis. This dissertation focuses on the design and application of PADs, especially ones that integrate with electrochemical systems, to tackle various problems in analytical chemistry, such as multi-analyte separation, sample enrichment, and sensitive detection. Four types of PADs are described in this dissertation. The first PAD (oPAD-Ep) is designed for multi-analyte separation. The oPAD-Ep is fabricated using the principle of origami to create a stack of connected paper layers as an electrophoresis channel. Due to the thinness of paper, a high electric field can be achieved with low voltage supply. Serum proteins can be separated and the device can be unfolded for post-analysis. The second PAD (oPAD-ITP) is designed on a similar principle as the oPAD-Ep, but it is applied for sample enrichment. The major modification is to adjust electrolyte conditions to enable isotachophoretic enrichment of analytes. DNA with various lengths can be enriched within a few minutes, and can be collected on one of the paper folds. The third PAD (hyPAD) also focuses on sample enrichment. The device is assembled with two different paper materials, nitrocellulose and cellulose. The hyPAD can perform faradaic ion concentration polarization experiments. This technique uses faradaic electrochemistry to create a local electric field gradient in the paper channel and can enrich charged analytes including: DNA, proteins, and nanoparticles. The fourth PAD (oSlip-DNA) focuses on sensitive electrochemical detection of DNA hybridization assays. This method integrates magnetic enrichment and electrochemical signal amplification via silver nanoparticles. Using voltammetry, sensitive detection of Hepatitis B Virus DNA is achieved on the low-cost device.Chemistr