Gallionella ferruginea is an iron-oxidizing chemolithotrophic micro-organism that lives in low-oxygen conditions (0.1–1.5 mg L−1 saturation). It produces a stalk structure from the concave side of the cell depending on population development, pH and redox conditions. After Gallionella oxidizes ferrous iron, bacteriogenic iron oxides (BIOS) precipitate on the stalk material and over time the stalks and/or the precipitated BIOS attenuate trace metals from surrounding groundwater. Gallionella ferruginea biofilms were cultured in situ in an artificial channel (2000 × 300 × 250 mm) using groundwater sourced from a borehole 297 m below sea level in the Äspö Hard Rock Laboratory in southern Sweden. The pH of the groundwater in the channels was always between 7.4 and 7.7 with oxygen saturation below 1.5 mg L−1 and Eh between 100 and 200 mV. Oxygen eventually declined to <0.3 mg L−1, terminating prolific biofilm growth. Biofilms formed within 2 weeks and were sampled every 2 weeks over 3 months. Cell number, stalk length and ferric iron concentration were measured for each sample and trace metal concentration was measured by inductively coupled plasma mass spectrometry. Results from well-developed in situ biofilms suggest that Gallionella could concentrate metals at levels up to 1 × 103-fold higher than found within the host rock and more than 1 × 106 times the levels found in the groundwater. These new experiments were used to support the results from the well-developed biofilms and to relate biofilm development and population characteristics to metal attenuation. After 3 months, rare earth element (REE) plots indicated that BIOS can accumulate metals at levels up to 1 × 104-fold higher than found in the groundwater and fractionate heavy rare earth elements over light rare earth elements. Generally the presence of the organic phase promotes the adsorption of all lanthanides and actinides that are not adsorbed by the inorganic phase. The iron oxides are directly correlated with stalk length (R = 0.96), indicating that rapid REE and actinide adsorption requires both iron oxides and a nucleating biological structure for the iron oxides.