Ethanol as a non-toxic solvent can dissolve PMMA and form acrylate monomers which diffuse across the interface of two layers and results in strong bonding upon the exposure of UV light. The device needed to be kept at 120 ºC for 2 hours under 1 kg weight . Doung et al. applied the same method to make a hybrid PLA/PMMA microfluidic device 107 . They used a spin coater to uniformly spread ethanol onto the layers to prevent bubble formation. The bonding was performed at a low temperature of 55 ℃. In another study, Doung et al. sprayed ethanol onto PMMA and ABS layers for bonding . After bringing the layers in contact, the device was put under UV to generate the temperature (55 °C to 60 °C) needed for dissolving the surface layer of PMMA. They also performed a post-heat treatment to release the residual stress caused after UV exposure. In order to enhance the efficiency of solvent bonding method, grooves were embedded in Persson et al.'s design to retain solvent in the vicinity of microchannels. Using SciGrip 4 as the solvent, they managed to create strong bonding between PMMA microchannels sandwiching a PETE membrane (Figure ) . Solvent bonding is usually not compatible with pre-functionalization as well, since normally both top and bottom thermoplastic layers should be treated with the solvent, which can eliminate the functional groups and denature the immobilized biomolecules. This method is achievable without denaturing pre-immobilized biomolecules only if the lid polymer layer (without the immobilized biomolecules) is in contact with the solvent. Moreover, the solvent bonding should be done at room temperature to maintain the functionality of the biomolecules, and the transparency of the layers should not deteriorate. PMMA-PMMA bonding at room temperature could be done using ethyl acetate and isopropanol (35:65%v/v) together with the lamination technique . Furthermore, Keller et al. used 35% cyclohexane in acetone to dissolve a COC polymer layer causing it to become "tacky" . They chose a non-penetrating polar acetone solvent to inhibit the excessive diffusion of the non-polar cyclohexane solvent into COC which can result in bubble formation and lack of transparency. In the proposed protocol the COC with an array of pre-immobilized biotin was bonded with another COC lid layer that had become tacky. They proved that the bonding technique does not denature the biotin compared to thermal fusion bonding.