Production of Hepatitis-B vaccine using recombinant methods MCQs With Answer

Production of Hepatitis-B vaccine using recombinant methods is a multidisciplinary process that transforms molecular cloning, host selection and bioprocess engineering into safe HBsAg-based vaccines. For B. Pharm students this topic covers recombinant DNA techniques, expression systems (yeast, Pichia, E. coli, mammalian), vector and promoter design, codon optimization, viral-like particle (VLP) assembly, downstream purification (chromatography, ultrafiltration), adjuvant formulation and key quality control assays (ELISA, potency, sterility). Understanding scale-up, glycosylation effects, antigen stability and regulatory standards is essential for vaccine development and manufacturing. This concise, keyword-rich overview links theory to practical production and analytical testing. Now let’s test your knowledge with 30 MCQs on this topic.

Q1. What is the primary antigen used in recombinant Hepatitis-B vaccines?

• Hepatitis B core antigen (HBcAg)
• Hepatitis B surface antigen (HBsAg)
• Hepatitis B e antigen (HBeAg)
• Hepatitis B polymerase

Correct Answer: Hepatitis B surface antigen (HBsAg)

Q2. Which expression host is most commonly used for licensed recombinant HBsAg vaccines?

• Escherichia coli
• Saccharomyces cerevisiae (yeast)
• CHO mammalian cells
• Pseudomonas aeruginosa

Correct Answer: Saccharomyces cerevisiae (yeast)

Q3. Why are viral-like particles (VLPs) important in recombinant Hepatitis-B vaccines?

• They contain viral DNA to stimulate T cells
• They mimic virus structure to enhance immunogenicity without infectious genome
• They increase vaccine viscosity
• They act as preservatives

Correct Answer: They mimic virus structure to enhance immunogenicity without infectious genome

Q4. Which promoter is commonly used in Pichia pastoris for high-level induction of recombinant proteins?

• GAL1 promoter
• T7 promoter
• AOX1 promoter
• Cytomegalovirus (CMV) promoter

Correct Answer: AOX1 promoter

Q5. What is the main reason E. coli is less preferred for HBsAg production?

• Low growth rate
• Inability to form disulfide bonds and glycosylation often leading to misfolding and inclusion bodies
• Lack of plasmid vectors
• High cost compared to yeast

Correct Answer: Inability to form disulfide bonds and glycosylation often leading to misfolding and inclusion bodies

Q6. Codon optimization in recombinant vaccine production primarily aims to:

• Reduce protein molecular weight
• Improve mRNA stability and translation efficiency in the chosen host
• Change antigen epitope specificity
• Eliminate post-translational modifications

Correct Answer: Improve mRNA stability and translation efficiency in the chosen host

Q7. Which secretion signal is widely used to direct HBsAg into the secretory pathway of yeast?

• PelB signal peptide
• Alpha mating factor (α-factor) prepro leader
• lac operon leader
• Type III secretion signal

Correct Answer: Alpha mating factor (α-factor) prepro leader

Q8. Which downstream technique is most effective for initial clarification after yeast fermentation?

• Size-exclusion chromatography
• Centrifugation and depth filtration
• Reverse-phase HPLC
• Immunoprecipitation

Correct Answer: Centrifugation and depth filtration

Q9. Which chromatographic method is commonly used for HBsAg purification based on charge differences?

• Affinity chromatography using protein A
• Ion exchange chromatography
• Reverse-phase chromatography
• Gel filtration using Sephadex

Correct Answer: Ion exchange chromatography

Q10. Aluminum-based adjuvants used in Hepatitis-B vaccines primarily function by:

• Directly neutralizing the virus
• Enhancing antigen uptake and stimulating local inflammation to boost immune response
• Acting as a stabilizer for HBsAg
• Providing nutrients for immune cells

Correct Answer: Enhancing antigen uptake and stimulating local inflammation to boost immune response

Q11. Which analytical assay is standard for quantifying HBsAg antigen content in vaccine batches?

• ELISA (enzyme-linked immunosorbent assay)
• qPCR for viral genome
• Gram staining
• Colorimetric protein assay only

Correct Answer: ELISA (enzyme-linked immunosorbent assay)

Q12. Glycosylation of HBsAg affects vaccine production because it:

• Eliminates antigenicity
• Affects protein folding, immunogenicity and recognition by host immune system
• Makes antigen water-insoluble
• Prevents VLP formation

Correct Answer: Affects protein folding, immunogenicity and recognition by host immune system

Q13. During scale-up of fermentation, one critical parameter to monitor for consistent HBsAg yield is:

• Agitation and dissolved oxygen
• Color of medium
• Odor intensity
• Volume of headspace only

Correct Answer: Agitation and dissolved oxygen

Q14. What is the purpose of ultrafiltration in downstream processing of recombinant HBsAg?

• To change the antigen sequence
• Concentration and buffer exchange of the antigen
• To sterilize by removing bacteria completely
• To degrade nucleic acids

Correct Answer: Concentration and buffer exchange of the antigen

Q15. Which test assesses residual host cell DNA or RNA in vaccine preparations?

• ELISA for HBsAg
• qPCR or nucleic acid quantification assays
• SDS-PAGE alone
• Endotoxin assay

Correct Answer: qPCR or nucleic acid quantification assays

Q16. Inclusion bodies formation indicates in recombinant expression that:

• Proteins are secreted efficiently
• High-level expression with misfolded aggregated protein often requiring refolding
• There is no expression
• The protein is fully glycosylated

Correct Answer: High-level expression with misfolded aggregated protein often requiring refolding

Q17. Which storage condition is typically recommended for final vialled recombinant Hepatitis-B vaccines?

• Room temperature for 6 months
• Frozen at -80°C only
• Refrigerated at 2–8°C
• Exposed to sunlight for stabilization

Correct Answer: Refrigerated at 2–8°C

Q18. Which assay is used to measure neutralizing antibody response after vaccination?

• Hemagglutination assay
• Neutralization assay or competitive ELISA measuring anti-HBs titers
• Total protein assay
• Gram staining

Correct Answer: Neutralization assay or competitive ELISA measuring anti-HBs titers

Q19. What is a common safety concern specific to recombinant vaccine production in bacterial hosts?

• Endotoxin (lipopolysaccharide) contamination
• Excess glycosylation
• Viral replication in product
• Aluminum toxicity from host cells

Correct Answer: Endotoxin (lipopolysaccharide) contamination

Q20. VLP assembly for HBsAg in yeast typically requires:

• Co-expression of viral polymerase
• Correct folding and oligomerization of HBsAg subunits
• Presence of viral genome
• Complete glycan removal

Correct Answer: Correct folding and oligomerization of HBsAg subunits

Q21. Which quality attribute ensures a vaccine batch is free from live microbial contaminants?

• Potency assay
• Sterility testing
• pH measurement
• Viscosity testing

Correct Answer: Sterility testing

Q22. WHO prequalification of a Hepatitis-B vaccine primarily evaluates:

• Color and label design
• Safety, efficacy, manufacturing quality and regulatory compliance
• Only cost of production
• Only animal testing results

Correct Answer: Safety, efficacy, manufacturing quality and regulatory compliance

Q23. Which method is most appropriate to detect HBsAg molecular weight and purity during development?

• SDS-PAGE and Western blot
• Gram staining
• Clinical chemistry analyzer
• Optical density at 600 nm only

Correct Answer: SDS-PAGE and Western blot

Q24. Why might Pichia pastoris be preferred over Saccharomyces cerevisiae for some recombinant protein productions?

• Pichia cannot glycosylate proteins
• Pichia often gives higher secreted yields and different glycosylation patterns suitable for some proteins
• Pichia grows only at low temperatures
• Pichia lacks fermentation protocols

Correct Answer: Pichia often gives higher secreted yields and different glycosylation patterns suitable for some proteins

Q25. During downstream purification, affinity tags are used mainly to:

• Increase cell growth rate
• Simplify capture step and increase purity of recombinant protein
• Remove endotoxin directly
• Denature the protein for analysis

Correct Answer: Simplify capture step and increase purity of recombinant protein

Q26. Which regulatory documentation is essential for a pharmaceutical manufacturer producing recombinant Hepatitis-B vaccine?

• Material Safety Data Sheet only
• Valid GMP records, batch release documentation and regulatory filings (e.g., dossier/CMC)
• Only marketing materials
• Only patent certificates

Correct Answer: Valid GMP records, batch release documentation and regulatory filings (e.g., dossier/CMC)

Q27. What is the role of buffer exchange after chromatographic steps?

• Add DNA contaminants
• Change formulation to the desired buffer and remove small molecule impurities
• Increase microbial contamination risk
• Reduce antigenicity intentionally

Correct Answer: Change formulation to the desired buffer and remove small molecule impurities

Q28. Which factor most directly influences VLP immunogenicity in recombinant Hepatitis-B vaccines?

• Color of the vaccine
• Native-like conformation and multivalent display of HBsAg epitopes
• Presence of viral RNA
• Manufacturing country

Correct Answer: Native-like conformation and multivalent display of HBsAg epitopes

Q29. Endotoxin removal is critical when producing vaccines in Gram-negative bacteria because endotoxin:

• Enhances vaccine potency and is always desirable
• Can cause strong pyrogenic responses and must be minimized
• Stabilizes proteins during storage
• Is required for VLP formation

Correct Answer: Can cause strong pyrogenic responses and must be minimized

Q30. Which stability study condition is most informative for real-time shelf-life prediction of a Hepatitis-B vaccine?

• Accelerated stability at very high temperatures only
• Real-time stability at recommended storage temperature (2–8°C) over intended shelf-life
• Short-term freeze-thaw cycles only
• Storage in sunlight

Correct Answer: Real-time stability at recommended storage temperature (2–8°C) over intended shelf-life

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