FIG 2.

Identification and isolation of single MPER-specific BM plasma cells by use of microengraving technology. (A) Diagram of the microengraving method. (B) Interrogation of slides after printing. MPER-specific Abs captured on MPER/liposome-coated slides were detected with Alexa 647-conjugated anti-mouse IgG. Human IL-6 was detected with Alexa 488-conjugated anti-human IL-6 to produce signals in every well for grid alignment and localization of MPER-specific signals. Human IL-6 was added to the printing medium. (C) Establishment of the microengraving system by using MPER-specific M1 hybridoma cells. The MPER-specific signals (green squares) were overlaid on the IL-6 signals (yellow squares) for localization of MPER-specific cells and retrieval for analysis. The MPER-specific M1 hybridoma cell line was used as an example. (D) Detection of MPER-specific plasma cells from bone marrow. Plasma cells were isolated from BM by depleting B220⁺ cells and CD49b⁺ cells and then enriching for CD138⁺ cells. A total of 2,674 MPER-specific signals were detected from 70,000 BM plasma cells isolated from one mouse immunized with Npalm-MPER/liposomes, whereas only 11 signals, all with signal intensities of <5,000, were detected from an unimmunized mouse.