Multiscreen htsip filter plates

Manufactured by Merck Group

The Multiscreen HTSIP Filter Plates are a laboratory filtration system designed for high-throughput screening applications. The plates feature a multi-well format and are used for rapid processing of multiple samples simultaneously.

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Lab products found in correlation

Chromogen 3 amino 9 ethylcarbazole, by Merck Group (1 mentions) Ionomycin, by Merck Group (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) Np osu, by Biosearch Technologies (1 mentions) Instantblue ultrafast protein stain, by Merck Group (1 mentions)

2 protocols using multiscreen htsip filter plates

Murine IFNγ ELISPOT Assay Protocol

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Mouse IFNγ ELISPOT assays were performed using the Ready-SET-Go! kit (eBioscience) on Multiscreen HTSIP Filter Plates (EMD Millipore). Briefly, plates were coated with anti-IFNγ capture antibody overnight and then blocked for 2 hours at 37°C with RPMI in 20% FBS. Fifty thousand negatively sorted CD8 + T cells were combined with 150,000 CD8- cells (flow-through) from vaccinated mice. Cells were incubated at 37°C for 24 hours with 5ug/mL of pooled peptides or 10 ug/mL of a single peptide. Positive control wells stimulated with phorbol 12-myristate 13-acetate (PMA) (25ng/mL) and Ionomycin (500ug/mL) (Sigma) and negative control DMSO-only (Sigma) wells were included on each plate. After overnight stimulation, the wells were decanted and incubated with biotinylated anti-mouse IFNγ primary antibody for 2 hours at 37°C, followed by Streptavidin-HRP antibody for 45 minutes. Spots were developed using 3-Amino-9-ethylcarbazole chromogen (Sigma) and washed extensively with tap water. After drying, plates were imaged and counted with the ImmunoSpot Analyzer (CTL, Cleveland, OH).

DeVette C.I., Gundlapalli H., Lai S.C., McMurtrey C.P., Hoover A.R., Gurung H.R., Chen W.R., Welm A.L, & Hildebrand W.H. (2019). A pipeline for identification and validation of tumor-specific antigens in a mouse model of metastatic breast cancer. Oncoimmunology, 9(1), 1685300.

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ELISA and ELISPOT Protocols for Protein Analysis

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Enzyme-linked immunosorbent assays (ELISAs) were done as described (48 (link)) with addition of 0.05% Tween 20 in block and wash buffers. 4-Hydroxy-3-nitrophenylacetyl hapten (NP) conjugated to BSA (NP-BSA, Ratio 28) was generated in-house with BSA fraction V (Roth Cat# 8076.3) and NP-OSu (Biosearch Technologies Cat# N1010-100). Plates were coated with 2 µg/ml NP-BSA or 1 µg/ml anti-light chain antibodies and developed with 1 µg/ml anti-isotype antibodies and the standards listed in Supplementary Table 1. For enzyme-linked immuno spot (ELISPOT) assays MultiScreen_HTS IP Filter Plates (Merck Cat# MSIPS4510) were coated and developed as described above for the ELISA plates, incubated with cells overnight, washed with 0.1% Tween 20 and processed according to the manufacturer’s instructions. For serum protein electrophoresis or immunofixation 10 µl serum was run on buffered agarose gels, pH8.6 Hydragel PROTEIN(E) (Sebia Cat# PN4100) or pH9.2 DOUBLE IF K20 (Sebia Cat# PN3036), and processed according to the manufacturer’s instructions. For proteomics, serum samples were run on multiple lanes of pH8.6 agarose gels and stained with InstantBlue Ultrafast Protein Stain (Sigma Cat# ISB1L). Excised bands were processed and analyzed by tandem mass spectrometry as described below.

Schmidt K., Sack U., Graf R., Winkler W., Popp O., Mertins P., Sommermann T., Kocks C, & Rajewsky K. (2020). B-Cell-Specific Myd88 L252P Expression Causes a Premalignant Gammopathy Resembling IgM MGUS. Frontiers in Immunology, 11, 602868.

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