This study investigated the influence of growth rate and glass slide preparation on bacterial attachment and biofilm development over time for methane-forming, anaerobic, mixed, microbial cultures. Photomicrographs and microscopic observations were also recorded. An anaerobic attachment vessel was designed, constructed, and used to quantify and visualize the initial attachment and biofilm development of chemostat grown bacterial cultures. The bacteria attached rapidly to washed/autoclaved glass slides. Within one to three hours, the number of irreversibly attached bacteria increased by approximately two orders of magnitude from 0 to 100-250 bacteria per 10,000 square micrometers. Only a slow increase in the number of attached bacteria was measured after the initial rapid increase. The counts of total bacteria after one week of inoculation were in a range of 250 to 450 bacteria per 10,000 square micrometers. No statistically significant difference was noted in the pattern of attachment for 8 days solids retention time (SRT) and 20 day SRT cultures. Two mathematical models were developed to describe the results. A significant percentage, usually 25%-75%, of the bacteria counted on the washed/autoclaved slides were methanogens. Final step autoclaving in the slide wash procedure had a statistically significant effect on attachment. Irreversibly attached bacteria counts on washed/unautoclaved slides over time were one half to one and one half orders of magnitude lower than the corresponding counts for washed/autoclaved slides. Scanning electron microscopy showed some cells do, and some do not, possess conspicuous appendages or extracellular fibers which appear to be used for attachment. At long inoculation times, more extensive development of extracellular fibers was observed sometimes and more amorphous, extracellular, gluelike material was present. Occasionally, extracellular fibers were observed to branch at longer inoculation times. Tip growth was proposed to account for this observation. At short and long inoculation times, cells attached as individuals and in clumps. The clumps were covered and/or interspersed with the gluelike material. Some clumps and individual cells appeared to have a ring around them, perhaps the secretion of extracellular polymers or enzymes. Higher concentrations of attached bacteria were sometimes observed on surface irregularities.