Advantages of stereolithographic 3D printing in the fabrication of the Affiblot device for dot-blot assays

Abstract

In stereolithographic (SLA) 3D printing, objects are constructed by exposing layers of photocurable resin to UV light. It is a highly user-friendly fabrication method that opens a possibility for technology sharing through CAD file online libraries. Here, we present a prototyping procedure of a microfluidics-enhanced dot-blot device (Affiblot) designed for simple and inexpensive screening of affinity molecule characteristics (antibodies, oligonucleotides, cell receptors, etc.). The incorporation of microfluidic features makes sample processing user-friendly, less time-consuming, and less laborious, all performed completely on-device, distinguishing it from other dot-blot devices. Initially, the Affiblot device was fabricated using CNC machining, which required significant investment in manual post-processing and resulted in low reproducibility. Utilization of SLA 3D printing reduced the amount of manual post-processing, which significantly streamlined the prototyping process. Moreover, it enabled the fabrication of previously impossible features, including internal fluidic channels. While 3D printing of sub-millimeter microchannels usually requires custom-built printers, we were able to fabricate microfluidic features on a readily available commercial printer. Open microchannels in the size range 200-300 μm could be fabricated with reliable repeatability and sealed with a replaceable foil. Economic aspects of device fabrication are also discussed.

Keywords: 3D printing; Antibody; Dot-blot; Microfluidics; Prototyping.

Fig. 1

Comparison of the CNC-milled and the 3D-printed version: a the impossibility of fabrication of internal channels by CNC milling led to the necessity of the 3-and-2 joint collector design connected to external output tubing; b 3D printing allowed individual collection from each row into an internal output channel, the overall size of the device was reduced for 3D printing to accommodate the build platform size limitation and to economize resin consumption

Fig. 2

a The MSLA-printed walls had a very gradual profile, which led to severe leakage all across the drain system; b the LFS-printed channels shrank during curing but sufficiently retained the liquid; c–f LFS prints of the microchannels of different widths; c the original CAD model with a channel width specification of 200 μm; d product printed from the model in image c; the channel walls fused, which blocked the channel off; e product printed from CAD files with a channel width specification of 300 μm; although not as well-defined, the channel was operational; f the product printed from CAD files with a channel width specification of 350 μm; the channel width of this print finally roughly reached 200 μm

Fig. 3

Prototyping of the designs of the drain system; a the 3-and-2-row joint collector outlet (remnant of the CNC-milled design) with an internal output channel (setup: 4 IO, 1 EO); b individual collector channel outlets for reduced risk of cross-contamination (setup: 10 IO, 1 EO); c second external outlet to balance the vacuum applied to each well (setup: 10 IO, 2 EO); d detail of the 3D-printed drainage wells and micro- and collector channels

Fig. 4

a Comparison of the previous CNC-micromachined functionality and the new 3D-printed Affiblot device; an identical calibration procedure was performed on both devices simultaneously; spots of an HE4 antigen were deposited onto the membrane, and two dilutions of two different clones of anti-HE4 antibodies were added to form the immune complexes. b Box charts of the correlations of colorimetric signals to the concentration of the HE4 antigen in the CNC-machined and the 3D-printed device