Skip to main content

Bead Extraction and Heat Dissociation (BEHD) For Mitigation of Drug Interference

 Bead Extraction and Heat Dissociation (BEHD) For Mitigation of Drug Interference

BEHD is the method developed for mitigating drug interference. BEHD method utilizes heat denaturation for the dissociation of the ADA-drug complex present in the samples. The samples are subsequently incubated with the sample and ADA is allowed to bind with Biotin-Drug and captured using streptavidin coated magnetic beads. 

Next, the samples are eluted using weak acids such as acetic acid or glycine, neutralized, and assayed in the ADA or NAb assay. The heat denaturation steps become critical when the drugs that are used to capture ADAs are susceptible to harsher acid-base conditions. This BEHD is used for the detection of ADA for cytokine therapy. 

Protocol For BEHD:

i) Heat Denaturation for Dissociation for ADA-Drug Complex

50ul to 100 ul of the serum samples containing ADAs are treated with mild heat at 62 degrees Celsius for 1 hour in thermocycler and allow the dissociation of drug-ADA complex

ii) Binding of ADA to Biotin Drug

50 ug/ml of Biotin-drug prepared in PBS are added the heat-treated samples and incubated for allowing the ADA binding to Biotin-Drug

iii) Capture of ADA-Biotin Drug Complex

The ADA-Biotin Drug complex is captured by mixing the streptavidin-coated magnetic beads to the ADA-drug sample mixture and incubating overnight at 4 degrees Celsius

iv) Washing and Removal of Residual Drug

After incubation, the magnetic Bead is washed using a Kingfisher or magnetic plate washer to remove unbound drug

v) Acid Elution

Weak acid including glycine or acetic acid are widely used to dissociate ADA from Biotin drug. Thus obtained drug unbound ADAs tested in ADA/NAb detection assay. 



Comments

Popular posts from this blog

Carbohydrate Metabolism: MCQs and answers on Glycolysis & Gluconeogenesis

                                      MCQ on Glycolysis & Gluconeogenesis 1) Which of the following enzyme is not involved in galactose metabolism? a) Glucokinase b) Galactokinase c) Galactose-1-Phosphate Uridyl transferase d) UDP-Galactose 4- epimerase 2) Which of the following enzyme is defective in galactosemia (type I) - a fatal genetic disorder in infants? a) Glucokinase b) Galactokinase c) Galactose-1-Phosphate Uridyl transferase d) UDP-Galactose 4- epimerase 3) In the liver, the accumulation of which of the following metabolite attenuates the inhibitory of ATP on phosphofructokinase? a) Glucose-6-Phosphate b) Citrate c) Fructose-1,6-Bisphosphate d) Fructose-2,6-Bisphosphate 4) Mutation in which of the following enzymes leads to a glycogen storage disease known as "Tarui’s disease"? a) Glucokinase b) Phosphofructokinase c) Phosphoglucomutase d) Pyruvate Kinase 5) E...

MCQs and Answers on cultivation (culture/incubation), Isolation and Identification of microorganisms: Medical Microbiology

40 plus questions - Multiple Choice Questions on Classification, Culture, and Identification of the microorganisms 1. Which of the following microorganism has the cocci cell shapes and sizes arranged usually in tetrad structures? a)  Streptococcus pneumoniae b)  Staphylococcus aureus c)  Chlamydia trachomatis d)  Neisseria meningitidis 2. What are the different growth morphology and cell structures used for the classification of fungi? Select all the correct answers: a) Yeast b) Mold c) Mycelia d) Protozoa 3. Which of the following media is formulated with additional nutrients to support the growth of fastidious or nutritionally demanding bacteria that may not grow well on basic media? a) Differential media b) Enriched media c) Nutrient agar (media) d) Selective media 4. Which of the following metabolic characteristic is a distinguishing characteristic and identification of colonies of  E. coli ? a) Hydrogen sulfide formation b) Indole Formation c) Lactose fe...

Multiple Choice Questions (MCQs) on Diabetes Mellitus: Pathogenesis, Diagnosis and Treatment

                                        MCQs on Diabetes mellitus 1) Diabetes mellitus is a disorder characterized by hyperglycemia.  Which of the following is not the common characteristic features of type 2 diabetes mellitus ? a) Impaired insulin secretion b) Increased Insulin resistance  c) Diabetic ketoacidosis d) Excessive hepatic glucose production 2) Which of the following are the characteristic features of type 1 diabetes mellitus? a) Type 1 diabetes is caused by an absolute deficiency of insulin. b) Type 1 diabetes is associated with the autoimmune destruction of beta cells.  c) Patients with  uncontrolled type 1 diabetes present with diabetic ketoacidosis d) All of the above   3) Which of the following serum measurements are not used for the diagnosis of diabetes mellitus? a) Fasting blood glucose d) Postprandial blood glucose  c) Insulin ...