Microwave cell lysis for ADDA‐ELISA analysis of total microcystins and nodularins

Abstract

A fast and effective microwave treatment procedure was developed for the ADDA-enzyme-linked immunosorbent assay analysis of total extracellular and intracellular microcystins and nodularins in drinking water and ambient water. This microwave treatment procedure for a full batch of samples required 10-20 min versus the 3-4 hr typically needed for three freeze/thaw cycles. The obtained lowest-concentration minimum reporting level and method detection limit were 0.21 and 0.09 μg/L microcystin-LR (MC-LR) equivalents, respectively. Mean recoveries of 80%-92% with relative standard deviations of 4.2%-9.0% were obtained for all studied water matrices fortified at concentrations of 0.3-2.0 μg/L MC-LR. Compared with US Environmental Protection Agency Method 546, this alternate procedure provided more effective cell lysis and subsequently detected significantly higher concentrations of total microcystins and nodularins in a few water samples collected during harmful algal blooms, such as 61.0 versus 34.4 μg/L MC-LR equivalents in Lake Erie and 11.5 versus 9.1 μg/L MC-LR equivalents in an Indiana reservoir.

Comparative Assessment of Physical and Chemical Cyanobacteria Cell Lysis Methods for Total Microcystin-LR Analysis

“…Microwave for 3 to 5 min achieved adequate MC-LR recovery in most samples. This was relatively consistent with other studies that demonstrated 9 to 20 min of microwave enabled recovery of total MCs [ 15 , 18 ]. However, recovery of MC-LR by microwave for cells suspended in CRW was lower than that applied to cells in phosphate buffer, and recovery of MC-LR in the control CRW showed greater variation when compared with the other lysis methods.…”

“…Such variation from microwave should be addressed by using sealed containers (able to withstand pressure) to hold samples during microwave, though this practice can limit sample volume relative to other methods [ 18 ]. Additionally, microwave without allowing samples to boil can minimize sample loss (and potential associated error) [ 15 ]. Microwave for 10 to 20 min has been shown to be more effective for MC recovery than three freeze-thaw cycles in natural bloom samples, and microwave lysis does not degrade multiple MC congeners [ 15 ].…”

“…Additionally, microwave without allowing samples to boil can minimize sample loss (and potential associated error) [ 15 ]. Microwave for 10 to 20 min has been shown to be more effective for MC recovery than three freeze-thaw cycles in natural bloom samples, and microwave lysis does not degrade multiple MC congeners [ 15 ].…”

“…Within a broader category of “thermal” cell lysis, autoclaving and boiling cells have been shown to moderately disrupt colonies and cells [ 24 ]. Based on the successful MC recovery previously observed via microwave when boiling of samples was avoided, microwave is able to lyse cells despite not causing complete destruction of cell morphology [ 15 ].…”

“…A wide variety of cell lysis methods are employed in literature, including lyophilization, bead beating, microwave, autoclaving, boiling, freeze-thaw, sonication, and chemical lysis. While the freeze-thaw process is frequently considered a standard method to lyse cells [ 12 ], it can require more time than other methods and may not recover all intracellular cyanotoxins present in samples [ 13 , 14 , 15 ]. There is a need to standardize and validate alternative cell lysis methods used for quantifying total cyanotoxins in order to give water-quality providers and laboratories more options to meet their response time goals for total cyanotoxin analysis.…”