Toxicity is crucial in biological research and plays an important role in drug development. Toxicity studies aim to remove toxic compounds early in the drug development process, as drug-induced toxicities are the leading cause of drug failures.

Thermo Fisher Scientific offers sensitive and accurate assays to probe and understand the effects of cytotoxins in biological systems. A variety of soluble biomarkers are available in different formats to suit your research needs.

 

Overview of toxicology research and testing

Toxicology encompasses scientific disciplines such as biology, chemistry, pharmacology, and medicine. Covering the entire biological system from cells to ecosystems, toxicology is the study of adverse effects of compounds on living organisms. The effects of toxins are influenced through their ADME (absorption, distribution, metabolism, and excretion) properties, along with their interactions with other molecules or cellular components [1].

Toxic substances interact in vivo in four steps. First, toxins must reach the blood stream. They can be absorbed through the gastrointestinal tract, skin exposure, or via lung inhalation. Once absorbed, toxic substances then enter the circulatory system through lymph and/or blood and get distributed to various organs and tissues.

Circulating toxins undergo metabolism or biotransformation, where the compounds get chemically transformed into metabolites that may elicit a weaker or stronger toxicological effect in vivo. Finally, toxins and/or their metabolites are excreted from the body. If a toxin can be rapidly eliminated from the body, it will have less chance of accumulating at the target site and inflicting damage. The most common methods of excretion are through the renal system or the digestive tract [2].


General testing methods

Toxicology testing can be broadly grouped into different categories. In research settings, synthetic and natural products/compounds are tested to assess their potential toxicological effects. In development and manufacturing of regulated materials for sale, like medicines, pesticides, cosmetics, additives, etc., all compounds must undergo toxicity testing on their final products prior to public use.

Testing methods will uncover species, organ, and dose-specific effects of compounds in question. The most common methods for testing are studying accidental exposure, in vitro studies using cell lines, and in vivo studies involving animal models [3]. At the end of testing, the LD50 of the compound is determined, which gives a value known to be a lethal dosage in 50% of the tested population.


ELISA and bead-based testing

As part of testing, it is important to understand the mechanism of action of compounds and their potential cytotoxic effects that are elicited in the body. Immunotoxicity can be measured in vitro using applications such as ELISA or ProcartaPlex multiplex immunoassays. These assays can be used to test a variety of biomarkers in the body to determine how toxins affect biomarkers. To that extent, Table 1 highlights a few examples of published data on toxicology research and testing using these methods.

A benefit of bead-based assays is the ability to multiplex—allowing simultaneous testing of different biomarkers at one time. A cost-effective and time-saving application, it requires only a small volume of sample (which is advantageous in the case of restricted/limited samples). Researchers can gain a comprehensive understanding of different toxicological effects due to the vast number of biomarkers across different species.

Table 1. List of publications highlighting immunological-based assays for toxicology research.

ReferenceSummaryTarget biomarkers/Panels usedTesting method
Effects of active and passive smoking on salivary cytokines levels in rats: A pilot study Study of the effects of smoking on salivary cytokine levels in rats.IL-6, IL-12p70, IFN-γProcartaplex
The mycotoxin alternariol suppresses lipopolysaccharide-induced inflammation in THP-1 derived macrophages targeting the NF-κB signalling pathway Study of the immunosuppressive effects of CD molecules & proinflammatory cytokines of the mycotoxin alternariol (metabolite of black mold) in human cells.NF-kB, IL-8, IL-6, TNF-α , IL-10Procartaplex
A New Immortalized Human Alveolar Epithelial Cell Model to Study Lung Injury and Toxicity on a Breathing Lung-On-Chip System In vitro evaluation of inhalation toxicity in a distal alveolar region model using human cell line.IL-8ELISA
Pulmonary inflammatory response from co-exposure to LPS and glyphosate Assessment of the effects of agricultural respiratory toxins, glyphosate and endotoxin in mice and their resulting lung inflammatory responses.TNF-α, keratinocyte chemoattractant (KC), MCP-1, MIP-2, IL-1β, IL-10, IL-13, IL-33, IL-4, IL-5, and IL-6ProcartaPlex
Formaldehyde exposure induces differentiation of regulatory T cells via the NFAT‑mediated T cell receptor signalling pathway in Yucatan minipigs Investigation of formaldehyde exposure and resulting toxicological effects on minipigs.IL-4, IFN-γ, TNF-αProcartaPlex
Topical application of the quaternary ammonium compound Didecyldimethylammonium chloride activates type 2 innate lymphoid cells and initiates a mixed-type allergic responseStudy of the hypersensitivity immune responses in murine models upon dermal exposure of an industrial/commercial compound in murine models.Th1/Th2/Th9/Th17/Th22/Treg Cytokine 17-Plex Mouse ProcartaPlex PanelProcartaPlex
Effects of different surface modifying agents on the cytotoxic and antimicrobial properties of ZnO nanoparticles Evaluation of cytotoxic properties of surface-modified ZnO nanoparticles by investigating inflammatory cytokine production in human cells.Inflammation 20-Plex Human ProcartaPlex PanelProcartaPlex
Evaluation of IL-1 blockade as an adjunct to linezolid therapy for tuberculosis in mice and macaques Assessment of reduced hematopoietic toxicity in a non-human primate model by addition of alpha-IL-IR1 to linezolid regimen.Cytokine & Chemokine 30-Plex NHP ProcartaPlex PanelProcartaPlex
Combined toxicity of air pollutants related to e-waste on inflammatory cytokines linked with neurotransmitters and pediatric behavioral problems Study of the toxicological effects of e-waste inhalation in humans.IL-1β, IL-6, TNF-αProcartaPlex

Toxicology research ELISA kits

ELISA kits can be used to detect and measure a variety of biomarkers that may be activated in toxicological settings. These kits can be used with a variety of biological sources and enable researchers to study the possible effects of toxins on different pathways in vivo, in distinct species. 

Invitrogen ELISA kits for popular targets such as IL-6, IFN gamma, VEGF etc. are listed in Table 2. Comparison of VEGF Human ELISA Kit with other commercial alternative is shown in Figure 1.

Search ELISA kits

Learn more about ELISA kits and components

Popular toxicology research protein targets and ELISA performance data

Table 2. View our ELISA kits for the following popular targets:

plot of Invitrogen VEGF Human ELISA Kit and competitor ELISA

Figure 1. Representative data using Invitrogen Human VEGF ELISA and competitor ELISA. Human serum samples were tested using VEGF Human ELISA Kit and compared to a competitor ELISA. Data shows a correlation factor of R2=0.9677.

Toxicology Research ProQuantum High Sensitivity Immunoassays

ProQuantum immunoassays are highly sensitive, high-performance kits used to detect target proteins with limited volume and do not require any specialized instruments. These assays utilize proximity-based amplification technology and combine analyte specific high-affinity antibody-antigen binding with signal detection and amplification capabilities similar to qPCR to achieve a simple yet powerful next-generation protein quantitation platform.

Invitrogen ProQuantum immunoassay kits for popular targets such as IL-6, IL-18, TNF alpha etc. are listed in Table 3. Standard curve of IL-6 using IL-6 Human ProQuantum Immunoassay Kit is shown in Figure 2.

Search all ProQuantum assays

Learn more about how the ProQuantum immunoassays work ›

Read BioProbes Journal article: Introducing ProQuantum High-Sensitivity Immunoassays—The new generation of target-specific protein quantitation

Popular toxicology research protein targets and ProQuantum assay performance data

Table 3. Toxicology-related ProQuantum immunoassays. View our ProQuantum immunoassay kits for the following popular targets:

Dot-plot of increasing IL-6 concentrations

Figure 2. Representative standard curve of Human IL-6. The standard curve for IL-6 using IL-6 Human ProQuantum Immunoassay Kit shows a broad dynamic range (0.0064–5,000 pg/mL) of IL-6 protein.

Toxicology research ProcartaPlex Multiplex Immunoassays

Invitrogen ProcartaPlex panels allow concurrent detection and measurement of soluble biomarkers implicated with toxin administration. Select from our preconfigured panels that are specifically designed and optimized for hepatotoxicity and nephrotoxicity research (Table 4) or use the panel configurator to customize a panel suited to your research needs associated with toxicology research and testing.

ProcartaPlex assays for liver and kidney toxicity enable the simultaneous quantification of up to 11 toxicity targets (including FDA-recommended biomarkers) in a single well [4]. Among others, highly relevant CYP targets for ADME/Tox studies are part of the ProcartaPlex menu. These assays can be used, along with other assay types such as enzyme activity assays or QuantiGene Plex mRNA to obtain a more holistic picture of your drug candidate (Figure 3). Organ-specific toxicity tests, specifically for the kidney are of high interest because it is a central detoxification organ that is exposed to drugs, reactive metabolites, and environmental chemicals. Kidney toxicity is routinely assessed during preclinical safety evaluations. Nephrotoxicity assays are tested against the most common sample types such as urine, serum/plasma and cell culture supernatant.

ProcartaPlex Liver Toxicity Panel

ProcartaPlex Kidney Toxicity Panel

ProcartaPlex Liver toxicity assays are available as preconfigured panels that are optimized for use with cell lysates. For example, ProcartaPlex Liver Toxicity Panels and matched QuantiGene Plex mRNA assays can be used to measure the effect of commonly utilized CYP3A4 inducers, viz. Rifampicin, Phenobarbital, 3-Methylcholanthrene and Ritonavir for evaluation of CYP3A4 induction by mRNA, and protein expression in primary human hepatocytes (Figure 3).  The enzymatic activity was checked to complement this data set.ProcartaPlex Kidney Toxicity assays are offered in multiple formats including preconfigured panels, simplex kits and custom blended Mix & Match panels. Preconfigured ProcartaPlex Kidney Toxicity Panels can be used to quantify up to 11 key urinary proteins simultaneously in a single well. For example, urine samples from individuals suffering from kidney diseases (AKI, RKF and CKDu) and healthy donors (control) were tested using Procartaplex Kidney Toxicity Panels (Figure 4) and (Figure 5). A significant increase of renal protein markers was observed in urine samples of kidney disease individuals compared to healthy donors.

 

ProcartaPlex panel configurator

Learn more about ProcartaPlex multiplex immunoassays

Preconfigured toxicology related-multiplex immunoassay panel data

Graph showing enzyme activity, mRNA and protein expression of CYP3A4 after Rifampicin, Phenobarbital, 3-Methylcholanthrene and Ritonavir submission

Figure 3. Effect of Rifampicin, Phenobarbital, 3-Methylcholanthrene and Ritonavir on CYP3A4 induction. Cryopreserved primary human hepatocytes (0.5-0.7x 105/well) were cultured in 96-well plates and treated with Rifampicin (30 µM), Phenobarbital (2000 µM), 3-Methylcholanthrene (2 µM) or Ritonavir (10 µM) for 48h. CYP3A4 induction was assessed by protein expression, enzyme activity and mRNA analysis (A) in cell supernatants using LC-MS/MS, (B) in lysed primary human hepatocytes using the ProcartaPlex Human Liver Toxicity Panel 1, 6plex and matched QuantiGene Plex mRNA assays (C). Data is expressed as fold change over the vehicle control (0.1% DMSO) and normalized mRNA/GAPDH ratio and protein/GAPDH ratio. Threshold of 2-fold induction over the vehicle control is indicated by dashed lines.

 

Figure 4. Urinary protein markers in acute kidney injury (AKI) and renal kidney failure (RKF) individual samples. The ProcartaPlex Human Kidney Toxicity Panel 1, 11plex and ProcartaPlex Human Kidney Toxicity Panel 2, 9plex were used to test key urinary protein markers in individuals. Data shows significantly increased markers in urine samples of 10 Acute Kidney Injury (AKI) and 13 Renal Kidney Failure (RKF) individuals compared to controls (healthy donor).

 

Figure 5. Urinary protein markers in chronic kidney disease of uncertain etiology (CKDu) individuals. The urine samples from 40 individuals were tested using ProcartaPlex Human Kidney Toxicity Panel 1, 11plex and ProcartaPlex Human Kidney Toxicity Panel 2, 9plex. Data shows significant increase in 10 protein markers in CKDu individuals compared to controls (healthy donor).


Table 4. Preconfigured ProcartaPlex multiplex immunoassay panels for toxicology-related research.

Product nameSizeCat. No.

ProcartaPlex Human Liver Toxicity Panel 1, 6plex

Targets: CYP1A2, CYP3A4, CYP2B6, CYP2C9, Beta-2-microglobulin (B2M), GAPDH

96 TestsEPX060-15857-901

ProcartaPlex Human Liver Toxicity Panel 2, 5plex

Targets: CYP2D6, CYP2C19, CP2E1, Beta-2-microglobulin (B2M), GAPDH

96 TestsEPX050-15858-901

ProcartaPlex Human Kidney Toxicity Panel 1, 11plex

Target list [bead region]: KIM-1/HAVCR/TIM-1 (Kidney injury molecule 1), Renin, Calbindin, Osteoactivin/GPNMB (Transmembrane Glycoprotein NMB), GSTA1 (Glutathione S-transferase A1), IL-18, IP-10 (CXCL10), MCP-1 (CCL2), VEGF-A, Clusterin (APO-J), RBP4 (Retinol-binding protein 4)

96 testsEPX110-15855-901

ProcartaPlex Human Kidney Toxicity Panel 2, 9plex

Target list [bead region]: Uromodulin, Alpha-1-microglobulin, TFF3 (Trefoil factor 3), Osteopontin (OPN), Cystatin C, NGAL, Beta-2-microglobulin (B2M), TIMP-1, EGF

96 testsEPX090-15856-901

ProcartaPlex Rat Kidney Toxicity Panel 1, 5plex

Target list [bead region]: Calbindin, KIM-1, Osteopontin (OPN), TFF3, VEGF-A

96 testsEPX050-30124-901

ProcartaPlex Rat Kidney Toxicity Panel 2, 5plex

Target list [bead region]: Albumin, Cystatin, Clusterin (Apo-J), NGAL, TIMP-1

96 testsEPX050-30125-901


Multiplex gene expression and protein assays

QuantiGene RNA gene expression assays provide a fast and high-throughput solution for multiplexed gene expression quantitation, with simultaneous measurement of up to 80 genes of interest in a single well of a 96- or 384-well plate. The QuantiGene Plex assay is based on hybridization and incorporates branched DNA (bDNA) technology, which uses signal amplification rather than target amplification for direct measurement of RNA transcripts. The assay is run on the Luminex® platform, has a simple workflow and does not require RNA purification. These features allow the user to merge the QuantiGene workflow for gene expression profiling with the ProcartaPlex workflow for protein quantitation (Figure 6) using the same sample. To study the gene expression of relevant ADME (absorption, distribution, metabolism, and excretion) and toxicological biomarkers, the QuantiGene Plex Human ADMETOX Panel, 80-plex is available to analyze 80 genetic targets implicated in drug metabolism, including enzymes, transporters, and CYPs.

Learn more about QuantiGene RNA assays for gene expression profiling and ADME/DMPK/Toxicology testing

Figure 6. Combined workflow for QuantiGene gene expression and ProcartaPlex protein quantitation assays.

Overview of toxicology research and testing

Toxicology encompasses scientific disciplines such as biology, chemistry, pharmacology, and medicine. Covering the entire biological system from cells to ecosystems, toxicology is the study of adverse effects of compounds on living organisms. The effects of toxins are influenced through their ADME (absorption, distribution, metabolism, and excretion) properties, along with their interactions with other molecules or cellular components [1].

Toxic substances interact in vivo in four steps. First, toxins must reach the blood stream. They can be absorbed through the gastrointestinal tract, skin exposure, or via lung inhalation. Once absorbed, toxic substances then enter the circulatory system through lymph and/or blood and get distributed to various organs and tissues.

Circulating toxins undergo metabolism or biotransformation, where the compounds get chemically transformed into metabolites that may elicit a weaker or stronger toxicological effect in vivo. Finally, toxins and/or their metabolites are excreted from the body. If a toxin can be rapidly eliminated from the body, it will have less chance of accumulating at the target site and inflicting damage. The most common methods of excretion are through the renal system or the digestive tract [2].


General testing methods

Toxicology testing can be broadly grouped into different categories. In research settings, synthetic and natural products/compounds are tested to assess their potential toxicological effects. In development and manufacturing of regulated materials for sale, like medicines, pesticides, cosmetics, additives, etc., all compounds must undergo toxicity testing on their final products prior to public use.

Testing methods will uncover species, organ, and dose-specific effects of compounds in question. The most common methods for testing are studying accidental exposure, in vitro studies using cell lines, and in vivo studies involving animal models [3]. At the end of testing, the LD50 of the compound is determined, which gives a value known to be a lethal dosage in 50% of the tested population.


ELISA and bead-based testing

As part of testing, it is important to understand the mechanism of action of compounds and their potential cytotoxic effects that are elicited in the body. Immunotoxicity can be measured in vitro using applications such as ELISA or ProcartaPlex multiplex immunoassays. These assays can be used to test a variety of biomarkers in the body to determine how toxins affect biomarkers. To that extent, Table 1 highlights a few examples of published data on toxicology research and testing using these methods.

A benefit of bead-based assays is the ability to multiplex—allowing simultaneous testing of different biomarkers at one time. A cost-effective and time-saving application, it requires only a small volume of sample (which is advantageous in the case of restricted/limited samples). Researchers can gain a comprehensive understanding of different toxicological effects due to the vast number of biomarkers across different species.

Table 1. List of publications highlighting immunological-based assays for toxicology research.

ReferenceSummaryTarget biomarkers/Panels usedTesting method
Effects of active and passive smoking on salivary cytokines levels in rats: A pilot study Study of the effects of smoking on salivary cytokine levels in rats.IL-6, IL-12p70, IFN-γProcartaplex
The mycotoxin alternariol suppresses lipopolysaccharide-induced inflammation in THP-1 derived macrophages targeting the NF-κB signalling pathway Study of the immunosuppressive effects of CD molecules & proinflammatory cytokines of the mycotoxin alternariol (metabolite of black mold) in human cells.NF-kB, IL-8, IL-6, TNF-α , IL-10Procartaplex
A New Immortalized Human Alveolar Epithelial Cell Model to Study Lung Injury and Toxicity on a Breathing Lung-On-Chip System In vitro evaluation of inhalation toxicity in a distal alveolar region model using human cell line.IL-8ELISA
Pulmonary inflammatory response from co-exposure to LPS and glyphosate Assessment of the effects of agricultural respiratory toxins, glyphosate and endotoxin in mice and their resulting lung inflammatory responses.TNF-α, keratinocyte chemoattractant (KC), MCP-1, MIP-2, IL-1β, IL-10, IL-13, IL-33, IL-4, IL-5, and IL-6ProcartaPlex
Formaldehyde exposure induces differentiation of regulatory T cells via the NFAT‑mediated T cell receptor signalling pathway in Yucatan minipigs Investigation of formaldehyde exposure and resulting toxicological effects on minipigs.IL-4, IFN-γ, TNF-αProcartaPlex
Topical application of the quaternary ammonium compound Didecyldimethylammonium chloride activates type 2 innate lymphoid cells and initiates a mixed-type allergic responseStudy of the hypersensitivity immune responses in murine models upon dermal exposure of an industrial/commercial compound in murine models.Th1/Th2/Th9/Th17/Th22/Treg Cytokine 17-Plex Mouse ProcartaPlex PanelProcartaPlex
Effects of different surface modifying agents on the cytotoxic and antimicrobial properties of ZnO nanoparticles Evaluation of cytotoxic properties of surface-modified ZnO nanoparticles by investigating inflammatory cytokine production in human cells.Inflammation 20-Plex Human ProcartaPlex PanelProcartaPlex
Evaluation of IL-1 blockade as an adjunct to linezolid therapy for tuberculosis in mice and macaques Assessment of reduced hematopoietic toxicity in a non-human primate model by addition of alpha-IL-IR1 to linezolid regimen.Cytokine & Chemokine 30-Plex NHP ProcartaPlex PanelProcartaPlex
Combined toxicity of air pollutants related to e-waste on inflammatory cytokines linked with neurotransmitters and pediatric behavioral problems Study of the toxicological effects of e-waste inhalation in humans.IL-1β, IL-6, TNF-αProcartaPlex

Toxicology research ELISA kits

ELISA kits can be used to detect and measure a variety of biomarkers that may be activated in toxicological settings. These kits can be used with a variety of biological sources and enable researchers to study the possible effects of toxins on different pathways in vivo, in distinct species. 

Invitrogen ELISA kits for popular targets such as IL-6, IFN gamma, VEGF etc. are listed in Table 2. Comparison of VEGF Human ELISA Kit with other commercial alternative is shown in Figure 1.

Search ELISA kits

Learn more about ELISA kits and components

Popular toxicology research protein targets and ELISA performance data

Table 2. View our ELISA kits for the following popular targets:

plot of Invitrogen VEGF Human ELISA Kit and competitor ELISA

Figure 1. Representative data using Invitrogen Human VEGF ELISA and competitor ELISA. Human serum samples were tested using VEGF Human ELISA Kit and compared to a competitor ELISA. Data shows a correlation factor of R2=0.9677.

Toxicology Research ProQuantum High Sensitivity Immunoassays

ProQuantum immunoassays are highly sensitive, high-performance kits used to detect target proteins with limited volume and do not require any specialized instruments. These assays utilize proximity-based amplification technology and combine analyte specific high-affinity antibody-antigen binding with signal detection and amplification capabilities similar to qPCR to achieve a simple yet powerful next-generation protein quantitation platform.

Invitrogen ProQuantum immunoassay kits for popular targets such as IL-6, IL-18, TNF alpha etc. are listed in Table 3. Standard curve of IL-6 using IL-6 Human ProQuantum Immunoassay Kit is shown in Figure 2.

Search all ProQuantum assays

Learn more about how the ProQuantum immunoassays work ›

Read BioProbes Journal article: Introducing ProQuantum High-Sensitivity Immunoassays—The new generation of target-specific protein quantitation

Popular toxicology research protein targets and ProQuantum assay performance data

Table 3. Toxicology-related ProQuantum immunoassays. View our ProQuantum immunoassay kits for the following popular targets:

Dot-plot of increasing IL-6 concentrations

Figure 2. Representative standard curve of Human IL-6. The standard curve for IL-6 using IL-6 Human ProQuantum Immunoassay Kit shows a broad dynamic range (0.0064–5,000 pg/mL) of IL-6 protein.

Toxicology research ProcartaPlex Multiplex Immunoassays

Invitrogen ProcartaPlex panels allow concurrent detection and measurement of soluble biomarkers implicated with toxin administration. Select from our preconfigured panels that are specifically designed and optimized for hepatotoxicity and nephrotoxicity research (Table 4) or use the panel configurator to customize a panel suited to your research needs associated with toxicology research and testing.

ProcartaPlex assays for liver and kidney toxicity enable the simultaneous quantification of up to 11 toxicity targets (including FDA-recommended biomarkers) in a single well [4]. Among others, highly relevant CYP targets for ADME/Tox studies are part of the ProcartaPlex menu. These assays can be used, along with other assay types such as enzyme activity assays or QuantiGene Plex mRNA to obtain a more holistic picture of your drug candidate (Figure 3). Organ-specific toxicity tests, specifically for the kidney are of high interest because it is a central detoxification organ that is exposed to drugs, reactive metabolites, and environmental chemicals. Kidney toxicity is routinely assessed during preclinical safety evaluations. Nephrotoxicity assays are tested against the most common sample types such as urine, serum/plasma and cell culture supernatant.

ProcartaPlex Liver Toxicity Panel

ProcartaPlex Kidney Toxicity Panel

ProcartaPlex Liver toxicity assays are available as preconfigured panels that are optimized for use with cell lysates. For example, ProcartaPlex Liver Toxicity Panels and matched QuantiGene Plex mRNA assays can be used to measure the effect of commonly utilized CYP3A4 inducers, viz. Rifampicin, Phenobarbital, 3-Methylcholanthrene and Ritonavir for evaluation of CYP3A4 induction by mRNA, and protein expression in primary human hepatocytes (Figure 3).  The enzymatic activity was checked to complement this data set.ProcartaPlex Kidney Toxicity assays are offered in multiple formats including preconfigured panels, simplex kits and custom blended Mix & Match panels. Preconfigured ProcartaPlex Kidney Toxicity Panels can be used to quantify up to 11 key urinary proteins simultaneously in a single well. For example, urine samples from individuals suffering from kidney diseases (AKI, RKF and CKDu) and healthy donors (control) were tested using Procartaplex Kidney Toxicity Panels (Figure 4) and (Figure 5). A significant increase of renal protein markers was observed in urine samples of kidney disease individuals compared to healthy donors.

 

ProcartaPlex panel configurator

Learn more about ProcartaPlex multiplex immunoassays

Preconfigured toxicology related-multiplex immunoassay panel data

Graph showing enzyme activity, mRNA and protein expression of CYP3A4 after Rifampicin, Phenobarbital, 3-Methylcholanthrene and Ritonavir submission

Figure 3. Effect of Rifampicin, Phenobarbital, 3-Methylcholanthrene and Ritonavir on CYP3A4 induction. Cryopreserved primary human hepatocytes (0.5-0.7x 105/well) were cultured in 96-well plates and treated with Rifampicin (30 µM), Phenobarbital (2000 µM), 3-Methylcholanthrene (2 µM) or Ritonavir (10 µM) for 48h. CYP3A4 induction was assessed by protein expression, enzyme activity and mRNA analysis (A) in cell supernatants using LC-MS/MS, (B) in lysed primary human hepatocytes using the ProcartaPlex Human Liver Toxicity Panel 1, 6plex and matched QuantiGene Plex mRNA assays (C). Data is expressed as fold change over the vehicle control (0.1% DMSO) and normalized mRNA/GAPDH ratio and protein/GAPDH ratio. Threshold of 2-fold induction over the vehicle control is indicated by dashed lines.

 

Figure 4. Urinary protein markers in acute kidney injury (AKI) and renal kidney failure (RKF) individual samples. The ProcartaPlex Human Kidney Toxicity Panel 1, 11plex and ProcartaPlex Human Kidney Toxicity Panel 2, 9plex were used to test key urinary protein markers in individuals. Data shows significantly increased markers in urine samples of 10 Acute Kidney Injury (AKI) and 13 Renal Kidney Failure (RKF) individuals compared to controls (healthy donor).

 

Figure 5. Urinary protein markers in chronic kidney disease of uncertain etiology (CKDu) individuals. The urine samples from 40 individuals were tested using ProcartaPlex Human Kidney Toxicity Panel 1, 11plex and ProcartaPlex Human Kidney Toxicity Panel 2, 9plex. Data shows significant increase in 10 protein markers in CKDu individuals compared to controls (healthy donor).


Table 4. Preconfigured ProcartaPlex multiplex immunoassay panels for toxicology-related research.

Product nameSizeCat. No.

ProcartaPlex Human Liver Toxicity Panel 1, 6plex

Targets: CYP1A2, CYP3A4, CYP2B6, CYP2C9, Beta-2-microglobulin (B2M), GAPDH

96 TestsEPX060-15857-901

ProcartaPlex Human Liver Toxicity Panel 2, 5plex

Targets: CYP2D6, CYP2C19, CP2E1, Beta-2-microglobulin (B2M), GAPDH

96 TestsEPX050-15858-901

ProcartaPlex Human Kidney Toxicity Panel 1, 11plex

Target list [bead region]: KIM-1/HAVCR/TIM-1 (Kidney injury molecule 1), Renin, Calbindin, Osteoactivin/GPNMB (Transmembrane Glycoprotein NMB), GSTA1 (Glutathione S-transferase A1), IL-18, IP-10 (CXCL10), MCP-1 (CCL2), VEGF-A, Clusterin (APO-J), RBP4 (Retinol-binding protein 4)

96 testsEPX110-15855-901

ProcartaPlex Human Kidney Toxicity Panel 2, 9plex

Target list [bead region]: Uromodulin, Alpha-1-microglobulin, TFF3 (Trefoil factor 3), Osteopontin (OPN), Cystatin C, NGAL, Beta-2-microglobulin (B2M), TIMP-1, EGF

96 testsEPX090-15856-901

ProcartaPlex Rat Kidney Toxicity Panel 1, 5plex

Target list [bead region]: Calbindin, KIM-1, Osteopontin (OPN), TFF3, VEGF-A

96 testsEPX050-30124-901

ProcartaPlex Rat Kidney Toxicity Panel 2, 5plex

Target list [bead region]: Albumin, Cystatin, Clusterin (Apo-J), NGAL, TIMP-1

96 testsEPX050-30125-901


Multiplex gene expression and protein assays

QuantiGene RNA gene expression assays provide a fast and high-throughput solution for multiplexed gene expression quantitation, with simultaneous measurement of up to 80 genes of interest in a single well of a 96- or 384-well plate. The QuantiGene Plex assay is based on hybridization and incorporates branched DNA (bDNA) technology, which uses signal amplification rather than target amplification for direct measurement of RNA transcripts. The assay is run on the Luminex® platform, has a simple workflow and does not require RNA purification. These features allow the user to merge the QuantiGene workflow for gene expression profiling with the ProcartaPlex workflow for protein quantitation (Figure 6) using the same sample. To study the gene expression of relevant ADME (absorption, distribution, metabolism, and excretion) and toxicological biomarkers, the QuantiGene Plex Human ADMETOX Panel, 80-plex is available to analyze 80 genetic targets implicated in drug metabolism, including enzymes, transporters, and CYPs.

Learn more about QuantiGene RNA assays for gene expression profiling and ADME/DMPK/Toxicology testing

Figure 6. Combined workflow for QuantiGene gene expression and ProcartaPlex protein quantitation assays.

Additional resources for Toxicology Immunoassays

Immunoassay instruments

References

For Research Use Only. Not for use in diagnostic procedures.

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