ABSTRACT
Monoclonal antibody (mAb)-based enzyme-linked immunosorbent assay (ELISA) is a complementary diagnosis technique for neurocysticercosis (NCC), which detects circulating parasite antigen (Ag) indicative of viable infection and Ag levels that correlate well with the parasite burden. In this study, we compared the performance of two Ag-ELISA techniques for the detection of NCC. We assessed the agreement between our in-house TsW8/TsW5 Ag-ELISA and the widely used B158/B60 Ag-ELISA for measuring T. solium antigen levels in the sera from 113 patients with calcified, parenchymal, and subarachnoid NCC. Concordance was demonstrated evaluating the limits of agreement (LoAs) stratified by the type of NCC. Both ELISA's detected 47/48 (97.8%) subarachnoid NCC cases. In parenchymal and calcified NCC, the B158/B60 Ag-ELISA detected 19/24 (79.2%) and 18/41 (43.9%) cases, while the TsW8/TsW5 Ag-ELISA detected 21/24 (87.5%) and 13/41 (31.7%), respectively. Parenchymal and calcified NCC obtained a perfect agreement (100%), indicating that all sample results were within the predicted LoA, while for subarachnoid NCC, the agreement was 89.6%. The high concordance between the assays was confirmed by Lin's concordance coefficient (LCC = 0.97). Patients with viable parenchymal NCC (LCC = 0.95) obtained the highest concordance between assays, followed by subarachnoid NCC (LCC = 0.93) and calcified NCC (LCC = 0.92). The TsW8/TsW5 Ag-ELISA and B158/B60 Ag-ELISA showed high Ag measurement correlations across diverse types of NCC.
ABSTRACT
The use of L protein coupled magnetic particles for the concentration and purification of immunoglobulin M (mIgM) monoclonal antibodies against Taenia solium was evaluated. Three concentration methods and different elution times were evaluated and the ratio of particles to the ratio of mIgM was optimized. It is demonstrated that: 1) with the use of magnetic particles, a previous concentration of mIgM is not required, which reduces the manipulation of the antibodies and improves the recovery, 2) the use of a binding buffer can be omitted, since the pH of most cell culture supernatants are neutral, and 3) longer elution times (~ 45 minutes) are needed to increase recovery to a level greater than 80%. The study demonstrates that the use of L protein-coupled magnetic particles is a simple and efficient tool for mIgM concentration and purification.
Se evaluó el uso de partículas magnéticas acopladas a proteína L para la concentración y purificación de anticuerpos monoclonales inmunoglobulina M (mIgM) contra Taenia solium. Se evaluaron tres métodos de concentración y diferentes tiempos de elución y se optimizó la proporción de partículas a la proporción de mIgM. Demostramos que: 1) con el uso partículas magnéticas no se requiere de una concentración previa de mIgM, lo que disminuye la manipulación de los anticuerpos y mejora la recuperación, 2) se puede omitir el uso de un tampón de unión, ya que el pH de la mayoría de los sobrenadantes de cultivo celular son neutros, y 3) se necesitan tiempos de elución más largos (~45 minutos) para aumentar la recuperación a un nivel mayor a 80%. El estudio demuestra que el uso de partículas magnéticas acopladas a proteína L es una herramienta simple y eficiente para la concentración y purificación de mIgM.
Subject(s)
Antibodies, Monoclonal , Immunoglobulin M , Magnetic Phenomena , Taenia solium , Animals , Antibodies, Monoclonal/isolation & purification , Immunoglobulin M/immunology , Taenia solium/immunologyABSTRACT
RESUMEN Se evaluó el uso de partículas magnéticas acopladas a proteína L para la concentración y purificación de anticuerpos monoclonales inmunoglobulina M (mIgM) contra Taenia solium. Se evaluaron tres métodos de concentración y diferentes tiempos de elución y se optimizó la proporción de partículas a la proporción de mIgM. Demostramos que: 1) con el uso partículas magnéticas no se requiere de una concentración previa de mIgM, lo que disminuye la manipulación de los anticuerpos y mejora la recuperación, 2) se puede omitir el uso de un tampón de unión, ya que el pH de la mayoría de los sobrenadantes de cultivo celular son neutros, y 3) se necesitan tiempos de elución más largos (~45 minutos) para aumentar la recuperación a un nivel mayor a 80%. El estudio demuestra que el uso de partículas magnéticas acopladas a proteína L es una herramienta simple y eficiente para la concentración y purificación de mIgM.
ABSTRACT The use of L protein coupled magnetic particles for the concentration and purification of immunoglobulin M (mIgM) monoclonal antibodies against Taenia solium was evaluated. Three concentration methods and different elution times were evaluated and the ratio of particles to the ratio of mIgM was optimized. It is demonstrated that: 1) with the use of magnetic particles, a previous concentration of mIgM is not required, which reduces the manipulation of the antibodies and improves the recovery, 2) the use of a binding buffer can be omitted, since the pH of most cell culture supernatants are neutral, and 3) longer elution times (~ 45 minutes) are needed to increase recovery to a level greater than 80%. The study demonstrates that the use of L protein-coupled magnetic particles is a simple and efficient tool for mIgM concentration and purification.
