CYANOBACTERIAL TOXINS - Cyanobacteria - page 545

reaction of cyanobacterial toxins with antibodies raised against a particular toxin variant very much
depends on differences in the degree of concentration between variants.
Immunoassays are also combined with other enzyme-linked immunosorbent (ELISA) assays.
Monoclonal antibodies (Kfi r et al ., 1986) and polyclonal antibodies (Brooks and Codd, 1988) isolated
against MC-LA were found to be specifi c to MC-LA. MC-LR-polylysine or MC-LR-ethylenediamine
conjugated with bovine serum albumin was used to raise antibodies. Such antibodies raised after
4 weeks of immunization of rabbits were characterized in a radioimmunoassay (RIA), ELISA and
indirect competitive ELISA. Of the two antigens MC-LR-ethylenediamine conjugated with bovine
serum albumin proved to be better. RIA and ELISA revealed that the antibodies cross-reacted with
other variants such as MC-RR, MC-YR and nodularin but had low reactivities with MC-LY and MC-LA
(Chu et al ., 1989). Nagata et al . (1995) demonstrated protective effects of specifi c monoclonal antibodies
on MC-LR-induced cell damages as assessed by morphological changes, lactate dehydrogenase
release into medium and cell viability measured by using a tetrazolium dye in primary rat hepatocyte
cultures. Weller et al . (2001) developed a direct competitive immunoassay of broad specifi city that
could be accomplished in a shortest time of 2 h and with a detection limit of 50 ng L -1 . Indirect
immunoassay (12 h), antiimmune complex immunoassay (48 h), and immunoassay based on
anti-idiotype antibody (12 h) exhibited lower cross-reactivities whereas direct immunoassay had
high cross-reactivity for nodularin as well. The percentage cross reactivities ranged from 100% for
MC-LR to a maximum of 156% in case of [Asp 3 , Dhb 7 ]MC-RR with intermediate percentages of cross-
reactivities for MC-RR (114%), MC-LA (115), MC-YR (118%), MC-LY (122%), MC-WR (147%), MC-LF
(123%), MC-LW (104%), [Asp 3 ]MC-HtyR (101%), [Asp 3 ]MC-LR (114%) and nodularin (102%).
Specifi c mono- and polyclonal antibodies against MCs conjugated with N-methyldehydroalanine
that is chemically modifi ed (aminoethylation) have been developed. Such monoclonal antibodies
were used in ELISA assays that could detect MCs as low as 1 ng ml -1 . A good correlation between
ELISA assays and HPLC was established. This is suggested to be extremely sensitive analytical tool
for direct measurement of toxins in cyanobacteria and water samples (Mikhailov et al ., 2001).
Polyclonal antibodies raised against MC-LR when employed in ELISA-assays showed cross
reaction with MC-LR and other variants of MC (Chu et al ., 1990). On the other hand, monoclonal
antibodies produced against MC-LR showed cross reaction with a number of MC variants and
nodularins in an ELISA format (Usleber et al ., 1995). ELISA requires long analysis time and needs
appropriate training (McDermott et al ., 1995; Pyo et al ., 2005). Fluorescence immunochromatography
method and HPLC have been used for detection of MCs. The advantages of the former method
are very short detection time and low detection limit while the latter method has an advantage of
quantifi cation of individual variants of MCs (Pyo et al ., 2005). Likewise, ELISA and HPLC have been
compared for the detection of MCs in environmental samples. Commercially available ELISA kit
was modifi ed suitably for quantitative determination of MCs within 2 to 3 days of sampling and
the assays were comparable to HPLC analysis. Modifi ed ELISA is more simple and highly sensitive
approach. Five species of Phormidium, i.e. P . bijugatum , P . molle , P . papyraceum , P . uncinatum and
P . autumnale hitherto not listed amongst toxic cyanobacteria caused neuro/hepatotoxic symptoms
in mice. Of the fi ve species, P . bijugatum extracts even caused death. The toxicity of the Phormidium
spp. was ascribed to STXs and MCs as confi rmed by ELISA (Teneva et al ., 2005b). The analysis of
serum samples of patients exposed to MCs during 1996 outbreak in Brazil by ELISA (19.9 ng ml -1 )
was almost comparable to the detection of these by LC/MS (21.2ng ml -1 ). This suggests that ELISA is
a simple, low-cost, accessible method to screen human serum for evidence of MC exposure (Hilborn
et al ., 2005).