Separation and detection of individual Aβ aggregates by capillary electrophoresis with laser-induced fluorescence detection

Advances and Pitfalls in the Capillary Electrophoresis Analysis of Aggregates of Beta Amyloid Peptides

“…As compared to previous CE-UV [23] or CE-LIF [24,25] studies that did not consider blanks (i.e., incubated or un-incubated matrix), the present work has revealed factors leading to interfering spikes, i.e., spikes related to material other than Aβ aggregates. Clearly, important causes of interfering spikes may be silica particles released from the capillary inlet in response to the mechanical stress to capillary occurring during the injection step and/or tiny dusts ripped out from the rubber stopper of vials.…”

“…In addition, the present work highlights the importance of running buffer nature and pH and suggests, for improving the performances of the analysis, the use of slightly acidic citrate or acetate buffer instead of neutral or slightly basic buffer as in previous studies [24,25]. In addition, care in the handling of running buffer has been emphasized when describing the limited stability of thioflavine T in such buffers, an effect that has been overlooked so far.…”

“…As it was not possible to have access to an electronic microscopy platform when performing the present study, the presence of fibrillar aggregates was not confirmed using an imaging approach. However, one can safely assume that our samples contained fibrils of Aβ peptides, because the incubation conditions we used (buffer, temperature, duration) are well known to lead to Aβ aggregates, some of them being fibrils [15,[23][24][25].…”

“…As compared to a previous study [25], it has the advantage to use a commercially available LIF instead of a home-made LIFA detector and to allow the direct analysis of sample, a buffer exchange step of sample prior to CE separation being unnecessary. Its selectivity mainly arises from the careful elimination of factors leading to interfering spikes and the optimization of thioflavine T containing running buffer.…”

“…In a first study, using a pH 9.5 running buffer, Kato et al [24] reported, the detection of a single spike in a sample of Aβ 1-42 was incubated for periods up to 6 days. In a second study [25], using a neutral (pH 7.79) buffer, CE was coupled to laser-induced fluorescence anisotropy (LIFA) detection for the separation, of spikes obtained after incubation of Aβ 1-40 or 1-42 for periods up to 7 days.…”