Detection Method
Indirect-ELISA
Intended Use
This kit is for research use only. Under no circumstances should it be used for therapeutic or human diagnostic applications.
Contents of Kit
| No. | Components | Size | Storage Conditions |
| 1 | PEG-BSA Coated Plate | 12×8 wells | -20°C |
| 2 | Anti-IgM HRP Stock (lyophilized) | 1 | -20°C |
| 3 | Reference Stock (lyophilized) | 1 | -20°C |
| 4 | 20× HRP PEG Wash | 50 ml | 2-8°C |
| 5 | HRP PEG Diluent | 50 ml | 2-8°C |
| 6 | TMB | 11 ml | 2-8°C |
| 7 | Stop Solution | 11 ml | 2-8°C |
Storage
The reference stock, HRP conjugate and the PEG-BSA coated plate should be stored at -20°C. All remaining kit components should be stored at 4°C. The microtiter plate should be kept in a sealed bag with desiccant. Kits will remain stable for six months from the date of purchase provided that the components are stored as described.
Performance Characteristics
Parallelism: To assess performance of the assay, two samples sample containing anti-PEG IgM at concentrations of 1226 and 2895 u/ml were serially diluted to produce values within the dynamic range of the assay.
General Description
Attachment of polyethylene glycol (PEG) chains to therapeutic biologic agents, a process referred to as PEGylation, prolongs the circulating half-life of the modified protein by slowing proteolytic degradation and by masking it from the immune system. However, it has been reported that repeated injections of PEGylated proteins can induce anti-PEG antibodies that increase the rate of clearance and decrease drug efficacy (accelerated blood clearance, or ABC phenomenon). To aid research in this key area, we have developed a human anti-PEG IgM ELISA kit.
Standard Curve
A typical standard curve with optical density readings at 450nm on the Y-axis against anti-PEG IgM concentrations on the X-axis is shown below. This curve is for the purpose of illustration only and should not be used to calculate unknowns.
Citations
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Polyethylene glycol (PEG) is a widely used polymer in the pharmaceutical industry for modifying therapeutic biologic agents, such as proteins and peptides. PEGylation involves attaching PEG chains to the surface of these molecules, which can enhance their pharmacokinetic properties, including prolonged circulation time and reduced susceptibility to enzymatic degradation. However, repeated exposure to PEGylated proteins can induce an immune response in some individuals, leading to the production of anti-PEG antibodies.
The presence of anti-PEG antibodies can have significant implications for the efficacy and safety of PEGylated therapeutics. When anti-PEG antibodies are present in the bloodstream, they can form immune complexes with PEGylated proteins, triggering an immune response. This immune response can result in a phenomenon known as the accelerated blood clearance (ABC) effect. The development of anti-PEG antibodies and the subsequent ABC phenomenon can vary among individuals and depend on factors such as the frequency and dosage of PEGylated protein administration, as well as the individual's immune characteristics. Pre-existing antibodies to PEG, which can be found in a subset of the population, may increase the likelihood of developing anti-PEG antibodies upon exposure to PEGylated therapeutics.
To mitigate the potential impact of anti-PEG antibodies on therapeutic efficacy, researchers are actively exploring strategies to minimize immunogenicity and reduce the risk of developing anti-PEG antibodies. This includes modifying the PEGylation process, such as using different PEG chain lengths or altering the attachment site on the protein, to minimize the immunogenicity of PEGylated proteins. Additionally, researchers are investigating alternative polymer-based modifications and exploring novel strategies, such as the use of stealth nanoparticles, to improve the pharmacokinetic properties of biologic agents without eliciting an immune response.
Alternative Names
Human Anti-PEG IgM ELISA
Human Anti-Polyetheylene Glycol IgM ELISA
Human Anti-Polyetheylene Glycol IgM ELISA Kit
Anti-PEG antibodies in the clinic: Current issues and beyond PEGylation
Journal of Controlled Release
Authors: Zhang, P., Sun, F., Liu, S., Jiang, S.
Abstract
The technique of attaching the polymer polyethylene glycol (PEG), or PEGylation, has brought more than ten protein drugs into market. The surface conjugation of PEG on proteins prolongs their blood circulation time and reduces immunogenicity by increasing their hydrodynamic size and masking surface epitopes. Despite this success, an emerging body of literature highlights the presence of antibodies produced by the immune system that specifically recognize and bind to PEG (anti-PEG Abs), including both pre-existing and treatment-induced Abs. More importantly, the existence of anti-PEG Abs has been correlated with loss of therapeutic efficacy and increase in adverse effects in several clinical reports examining different PEGylated therapeutics. To better understand the nature of anti-PEG immunity, we summarize a number of clinical reports and some critical animal studies regarding pre-existing and treatment-induced anti-PEG Abs. Various anti-PEG detection methods used in different studies were provided. Several protein modification technologies beyond PEGylation were also highlighted.
Anti-PEG immunity: emergence, characteristics, and unaddressed questions
Wiley Interdisciplinary Reviews: Nanomedicine and Nanobiotechnology
Authors: Yang Q, Lai S K.
Abstract
The modification of protein and nanoparticle therapeutics with polyethylene glycol (PEG), a flexible, uncharged, and highly hydrophilic polymer, is a widely adopted approach to reduce RES clearance, extend circulation time, and improve drug efficacy. Nevertheless, an emerging body of literature, generated by numerous research groups, demonstrates that the immune system can produce antibodies that specifically bind PEG, which can lead to the 'accelerated blood clearance' of PEGylated therapeutics. In animals, anti-PEG immunity is typically robust but short-lived and consists of a predominantly anti-PEG IgM response. Rodent studies suggest that the induction of anti-PEG antibodies (α-PEG Abs) primarily occurs through a type 2 T-cell independent mechanism. Although anti-PEG immunity is less well-studied in humans, the presence of α-PEG Abs has been correlated with reduced efficacy of PEGylated therapeutics in clinical trials. The prevalence of anti-PEG IgG and reports of memory immune responses, as well as the existence of α-PEG Abs in healthy untreated individuals, suggests that the mechanism(s) and features of human anti-PEG immune responses may differ from those of animal models. Many questions, including the incidence rate of pre-existing α-PEG Abs and immunological mechanism(s) of α-PEG Ab formation in humans, must be answered in order to fully address the potential complications of anti-PEG immunity.
COA
Certificate of Analysis
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