The COVID-19 pandemic set a new precedent for those working to get testing and medicines to the market as soon as possible. Discover how technology can help support your research of infectious disease cellular responses from detecting IgG in serum samples to identifying neutralizing antibodies in this new eBook.
In this eBook, you’ll learn about:
- Detecting SARS-CoV-2 IgG in serum samples with a luminescent immunoassay
- Qualitative measurement of SARS-CoV-2 spike and nucleocapsid IgG antibodies in serum samples
- Measuring viral-induced cytopathic effects during drug discovery and development
- Assessing virus neutralization with a rapid, HTS-friendly assay
- Identifying neutralizing antibodies with a surrogate virus neutralization test
Virology and Vaccine Research Microplate Reader Solutions | Molecular Devices
AND VACCINE RESEARCH:
From detecting IgG in serum samples to identifying neutralizing antibodies
We understand what a challenging time this is for those who are working to get accelerated testing and medicines to the market as soon as possible and are committed to supporting scientists that are researching cellular response to viruses for the development of therapeutics and vaccines.
Learn more about how our technology can help support your research of infectious disease cellular responses.
with a luminescent immunoassay 3
with a quantitative luminescence assay 4
dual reporter assay 5
in serum samples 6
IgG in cell line development 7
Low-volume nucleic acid quantitation 8
Measure total protein in cell lysates 9
HTS-friendly assay 10
As part of the immune response to SARS-CoV-2, infected individuals produce virus-specific antibodies that are detectable in the blood within one to three weeks after the onset of symptoms. Both immunoglobulin-G (IgG) and IgM appear almost simultaneously, with IgG remaining at a high level for
a longer time1, 2. The presence of detectable antibodies in serum resulting from infection or vaccination can protect against future infection, but the length of time these antibodies persist and the degree of protection they afford are among the many questions about SARS-CoV-2 that remain under intense investigation.
Scale up for HTS with a homogeneous,
Use a sample volume as little as 15 μL
Obtain results in less than one hour
Add Lumit™ Dx CoV-2 LgBiT and Lumit™ Dx CoV-2 SmBiT to plate wells.
Add samples and controls to plate wells. Incubate for 30 minutes.
Add Lumit™ Dx Detection Reagent. Incubate for 3 minutes.
Assay workflow for the Lumit Dx SARS-CoV-2 Immunoassay. The add-incubate-read format enables results in less than one hour and is amenable to automation and high-throughput screening.
Viral infection of mammalian cells often reduces cell viability and causes visible effects upon the cells such as changes in shape or size, or fusion with adjacent cells. These changes are called cytopathic effects (CPE), and they can be assessed using light microscopy or imaging systems, or measured using more quantitative means.
Easy mix-and-read method with results in just 10 minutes after reagent addition
Stable, sensitive luminescence readout suitable for screening
Generate data and analyze results automatically with SoftMax Pro Software
Add virus/compound, incubate
Add ATP detection reagent
1 2 3 4 5
Viral ToxGlo™ Assay workflow.
SpectraMax microplate reader
SoftMax Pro Software
Monitor NF-κB activation with a sensitive dual reporter assay on the SpectraMax iD5 microplate reader
Reporter gene assays are used to study the expression of eukaryotic genes. In dual reporter gene assays, cells are transfected with two vectors, the first containing an experimental reporter gene coupled to a regulated promoter of interest, and the second containing a control reporter gene coupled to a constitutive promoter. Normalizing the activity of the experimental reporter to the control reporter minimizes experimental variability.
Bioluminescent reporter systems using firefly and Renilla luciferases are widely used as co-reporters because the assays are easy to run and exquisitely sensitive. Dual-luciferase assays can elucidate a variety of cell
signaling pathways, including disruptions in signaling caused by viral infection.
Detect multiplexed luciferase assay results with a sensitive dual reporter assay
Improve accuracy of results by normalizing reporter expression to a control
Expedite results with a preconfigured SoftMax Pro Software protocol
NF-KB signaling pathway. TNF-α activates a signal cascade resulting in degradation of the NF-KB inhibitor IKB and release of NF-KB transcription factor to the nucleus.
Rapid, accurate, and frequent molecular testing is of critical importance in navigating the SARS-CoV-2 (COVID-19) pandemic. To advance our understanding of the relationship between infection and immunity, it is
critical to test not only for infection, but also for antibodies. The fundamental challenge is to identify and implement sensitive, consistent, and accessible testing methods that fulfill these requirements, all at an affordable price.
Gain >95% specificity for nucleocapsid and spike IgG antibodies
Measure up to 86 samples per kit with high-throughput, qualitative immunoassays
Acquire colorimetric readout with exceptional inter/
intra assay precision
Secondary antibody conjugate
SARS-CoV-2 IgG ELISA (indirect ELISA format). SARS-CoV-2 IgG ELISAs schematic. The bottom of the wells are coated with SARS-CoV-2 Nucleocapsid protein or Spike S1-receptor binding domain (antigen). If present in the serum sample, IgG specific for the antigen binds to the coated wells, and an
HRP-conjugated antibody enables detection of the bound IgG with the addition of substrate.
Calibrator concentration (ng/mL)
Calibrator curve for the SARS-CoV-2 Nucleocapsid IgG ELISA Kit. The curve was plotted in SoftMax Pro Software using a
4-parameter logistic curve fit. From this curve, results were calculated semi-quantitatively.
Biologic drugs are the largest and fastest growing segment of the pharmaceutical industry with sales of €500bn and an annual growth of 8%. Every manufacturing process for potential biologics begins with cell line development, whether it’s for clinical trials or a market launch.
Monoclonal antibodies (mAb) have established themselves as the leading biopharmaceutical therapeutic modality. The establishment of robust manufacturing platforms is key for antibody drug discovery efforts to seamlessly translate into clinical and commercial successes.
Fast, homogeneous, and automation-friendly assay with results for 96 samples in less than 15 minutes
Low sample volume and limited test sample pre-preparation, allowing analysis of samples in a crude matrix
Number of clones
Precise measurement of IgG from 2.5 to 2000 mg/L
Vector construction & transfection
Single cell isolation
Select/screen for productivity
Screen for quality
Screen for stability
The cell line development process from cell transfection to scale-up.
Nucleic acid quantitation is an essential assay that is part of many workflows in genetics, molecular, and cellular biology. The development of nucleic acid vaccines is yet another area where accurate quantitation of material at different steps in the process is critical. Various methods have been developed to quantitate nucleic acids, but the most commonly used technique is still ultra-violet (UV) spectrophotometry. The basis of spectrophotometry is that every molecule absorbs or transmits light over a certain wavelength range, and concentrations can be calculated by using the Beer- Lambert law with preceding knowledge of the sample’s molar extinction coefficient and the measurement pathlength.
Low-volume nucleic acid quantitation using as little as 2 µL of sample
High-throughput measurement of 24 or 64 samples at a time
Linear dynamic range of 3 decades for accurate quantitation without the need for dilution
low volume sample slide
SpectraDrop Microplate adapter
SpectraDrop Micro-Volume Microplate configuration. The low- volume sample slide has a mask delineating spots or ‘wells’ that hold 24 or 64 samples. The cover slide has 0.5-mm or 1.0-mm spacers for use with 2-µL or 4-µL samples, respectively.
DNA concentration (ng/μL)
DNA standard curves on the SpectraDrop plate, read on the SpectraMax Mini reader. Blue plot shows 2-µL samples; red plot shows 4-µL samples (r2 values were 0.991 and 1.000, respectively).
Quantitation of proteins in cell lysates is a key step for many downstream applications including western blots and enzyme linked immunosorbent assays (ELISAs), which are often used to identify viral proteins in clinical or research samples. The most popular protein quantitation assays are
colorimetric, relying on the change in color of a reagent in the presence of the protein of interest.
Here, we describe the use of the Pierce Rapid Gold BCA and Pierce Detergent Compatible Bradford assays to quantitate total protein in cell lysates.
Accurately assay protein concentrations with minimal incubation time
Obtain results quickly with the 8-channel read head
Plot standard curves and calculate sample protein concentrations automatically with preconfigured protocols in SoftMax Pro Software
Mean OD value
BSA standard curves for the BCA Rapid Gold assay (red plot, r2 = 1.000) and Detergent Compatible Bradford assay (blue plot, r2 = 0.999).
Rapid Gold BCA assay results. SoftMax Pro Software data table shows the calculated concentrations of serially diluted cell lysate.
Bradford assay results. SoftMax Pro Software data table shows calculated concentrations of cell lysate with the Detergent Compatible Bradford assay.
The worldwide COVID-19 pandemic caused by the SARS-CoV-2 virus has necessitated the fast-tracked development of many research tools for understanding this virus’s pathogenesis, as well as vaccine discovery and development. Assays for monitoring the immune response to infection by and vaccination against the virus are important to COVID-19 research.
Rapid, easy assay setup with an ELISA workflow
No BSL-3 or cell culture required
Results consistent with data from plaque reduction neutralization test
Cell and virus preparation
Add the above mixture to the wells pre-coated with ACE2 for 15 minutes
Incubate testing samples with virus for 1 hour
Add the above mixture to Vero-E6 cells and incubate for 1 hour
Incubate testing samples with HRP-RBD for 30 minutes
PRNT assay (BSL3 lab) sVNT kit (BSL2 lab)
Overlay 1% agarose and incubate for 2 days
Fix cells and stain
Add substrate and incubate for 15 minutes
Add s top solution and read
Total assay time = 1 hour
Total assay time = 2.5 days
Comparison of PRNT and sVNT kit workflows. The sVNT assay can be completed in one hour in a BSL-2 lab, while PRNT requires live SARS-CoV-2 virus and cumbersome cell culture techniques, taking more than two days at a BSL-3 lab.
1 PPDM Bioanalyitical Regulated Immunogenicity and Molecular, Merck and Co., Inc., Kenilworth, New Jersey; 2 Biologics Analytical Sciences, Merck and Co., Inc., Kenilworth, New Jersey; 3 PPDM Bioanalyitical Regulated Immunogenicity and Molecular, Merck and Co., Inc., West Point, Pennsylvania; 4 Biometrics Research, Merck and Co., Inc., West Point, Pennsylvania; 5 Q2 Solutions Vaccines, San Juan Capistrano, California
Vaccine immunogenicity and clinical efficacy are often assessed by the measure of serum-neutralizing antibodies. The present gold standard for detecting neutralizing antibodies against many viruses, including dengue, is the plaque/ focus reduction neutralization test (P/FRNT). The FRNT is a cell-based assay that inherits high variability, resulting in poor precision and has lengthy turnaround times. The virus reduction neutralization test (VRNT) is a high-throughput alternative to the standard low-throughput and laborious FRNT. The VRNT is similar to FRNT using unaltered wild-type virus and immunostaining, yet uses imaging cytometry to count virus-infected cells one day post-infection, reducing assay time and increasing overall throughput 15-fold. In addition, the VRNT has lowered variability relative to FRNT, which may be explained in part by the observation that foci overlap alters foci count and titer over time, in the FRNT.
The ability to count one infected cell, rather than waiting for overlapping foci to form, ensures accuracy and contributes to the precision (7–25% coefficient of variation) and sensitivity of the VRNT. Results from 81 clinical samples tested in the VRNT and FRNT show a clear positive relationship. During sample testing, a 96-well plate edge effect was noted and the elimination of this edge effect was achieved by a simple plate seeding technique. The VRNT is an improvement to the current neutralization assays for its shortened assay time, increased precision and throughput, and an alternative to the P/FRNT.
Am. J. Trop. Med. Hyg., 99(6), 2018, pp. 1430–1439 doi:10.4269/ajtmh.17-0948
Copyright © 2018 by The American Society of Tropical Medicine and Hygiene
Virus reduction neutralization test:
A single-cell imaging high-throughput virus neutralization assay for dengue
SpectraMax i3x Multi-Mode Microplate Reader
SpectraMax Mini Multi-Mode Microplate Reader
SpectraMax iD3 Multi-Mode Microplate Reader
SpectraMax M5 Multi-Mode Microplate Reader
SpectraMax iD5 Multi-Mode Microplate Reader
SpectraMax ABS/ABS Plus Absorbance Microplate Reader
Check our website for a current
USA and Canada +1.800.635.5577
United Kingdom +44.118.944.8000
Taiwan/Hong Kong +886.2.2656.7585
South Korea +82.2.3471.9531
listing of worldwide distributors. * Austria, Belgium, Denmark, Finland, France, Germany, Iceland, Ireland, Italy, Luxembourg, Netherlands, Portugal, Spain, Sweden, Switzerland and United Kingdom
The trademarks used herein are the property of Molecular Devices, LLC or their respective owners. Specifications subject to change without notice. Patents: www.moleculardevices.com/productpatents FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.
©2022 Molecular Devices, LLC
Printed in USA