Quantifying Tetanus Toxoid-Specific Antibody-Secreting Cells

TT-specific ASC were enumerated by ELISPOT, as previously described (15 (link), 19 (link), 20 (link), 38 (link)). MultiScreen High protein binding immobilon-P membrane plates (Millipore Corporation, Bedford, MA) were coated with 10 μg/ml TT overnight at 37°C, blocked with complete RPMI 1640 (Life Technologies BRL, Life Technologies, Paisley, U.K.). Duplicates of cells from spleen and bone marrow in four three-fold dilutions starting with 1 × 10⁷ cells in 100 μL in complete RPMI 1640 per well were incubated for 5 hours at 37°C, washed and incubated with ALP-goat anti-mouse IgG (Southern Biotechnology Associates) overnight at 4°C, and developed by 5-bromo-4-chloro-3-indolylphosphate and NBT in AP development buffer (Bio-Rad Labs, Hercules, CA). The number of spots, each representing a cell secreting TT-specific IgG antibodies, was counted with ELISPOT reader ImmunoSpot^® S6 ULTIMATE using ImmunoSpot^® SOFTWARE (Cellular Technology Limited (CTL) Europe, Bonn, Germany).

Free full text: Click here

Aradottir Pind A.A., Thorsdottir S., Magnusdottir G.J., Meinke A., Del Giudice G., Jonsdottir I, & Bjarnarson S.P. (2022). A comparative study of adjuvants effects on neonatal plasma cell survival niche in bone marrow and persistence of humoral immune responses. Frontiers in Immunology, 13, 904415.

Publication 2022

MultiScreen High protein binding immobilon-P membrane plates complete RPMI 1640 ALP-goat anti-mouse IgG 5-bromo-4-chloro-3-indolylphosphate ImmunoSpot® S6 ULTIMATE ImmunoSpot® SOFTWARE

Anti igg Bone marrow Buffer Cellular Dilutions Elispot Goat Igg antibodies Immobilon p Membrane Mouse Spleen Spots

Corresponding Organization : University of Iceland

Other organizations : Valneva (Austria), Toscana Life Sciences

Top 5 similar protocols

Variable analysis

independent variables

Dose of TT used to coat the ELISPOT plates (10 μg/ml)

dependent variables

Number of TT-specific IgG antibody-secreting cells in the spleen and bone marrow, as counted by the ELISPOT reader

control variables

Overnight coating of ELISPOT plates with TT at 37°C
Blocking of plates with complete RPMI 1640
Incubation of cell samples in complete RPMI 1640 for 5 hours at 37°C
Overnight incubation with ALP-goat anti-mouse IgG at 4°C
Development of ELISPOT plates using 5-bromo-4-chloro-3-indolylphosphate and NBT in AP development buffer

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

Based on most similar protocols

Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.

As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!