A new method for stirring thin liquid films has been developed and demonstrated to increase the sensitivity of immunofluorescence staining of polytene chromosomes. This liquid-on-liquid mixing (LOLM) technique uses a stirrer fluid, immiscible with the thin film, to transmit shear at the liquid-liquid interface. Here, we stir mineral oil layered over an aqueous thin film of antibody solution, which stains transcription apparatuses on chromosomes previously fixed to a glass slide. The quality of staining was assessed at varying antibody concentrations and incubation or stirring times. Our data indicate that the LOLM technique overcomes the diffusion barrier associated with traditional slide-based biological assays.
Bibliographical noteFunding Information:
The authors thank Janis Werner for offering advice on polytene chromosome preparation and staining; Eric Siggia for pointing out the possible diffusion bottleneck in slide-based assays; and Zsofia Franck, Steven J. Koch, Arthur La Porta, and Brian Schwartz for useful discussions. This publication was developed under the auspices of the Cornell University Center for Biotechnology, a New York State Center for Advanced Technology, supported by New York State and industrial sponsors. This material is based upon work supported in part by the STC Program of the National Science Foundation under Agreement No. ECS-9876771. This work made use of the computing facility of the Cornell Center for Materials Research (CCMR), supported through the National Science Foundation Materials Research Science and Engineering Centers (MRSEC) program (DMR-0079992).
- Immunofluorescence staining
- Polytene chromosomes
- Thin film