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USMLE Step 1 Quiz

USMLE Step 1 Quiz: Biochemical Laboratory Techniques

Practice Biochemical Laboratory Techniques in USMLE Step 1 with focused quiz questions that help you check what you know, review explanations, and build confidence with test-style prompts.

Question 1 / 20

0 of 20 answered

A 45-year-old man presents to a clinic for a routine health screening. He reports a history of intravenous drug use several years ago. A rapid screening test for HIV is positive. A confirmatory test is ordered, which involves separating viral components by size on a gel, transferring them to a membrane, and then using patient serum to detect antibodies that bind to these specific components.

This confirmatory test is designed to detect the presence of host antibodies against which of the following?

Select an answer to continue

What this quiz covers

This quiz focuses on Biochemical Laboratory Techniques, giving you a quick way to practice the rules, question types, and explanations that matter most for USMLE Step 1.

How to use this quiz

Try each quiz question before looking at the correct answer. Use the explanations to review missed ideas, then come back to similar questions until the pattern feels familiar.

All questions

Question 1

A 45-year-old man presents to a clinic for a routine health screening. He reports a history of intravenous drug use several years ago. A rapid screening test for HIV is positive. A confirmatory test is ordered, which involves separating viral components by size on a gel, transferring them to a membrane, and then using patient serum to detect antibodies that bind to these specific components.

This confirmatory test is designed to detect the presence of host antibodies against which of the following?

  1. Specific viral proteins (correct answer)
  2. Viral RNA sequences
  3. Integrated proviral DNA
  4. Viral reverse transcriptase enzyme activity

Explanation: The technique described is a Western blot, the standard confirmatory test for HIV. It works by detecting host antibodies (from patient serum) that bind to specific viral proteins (e.g., p24, gp41, gp120) that have been separated by gel electrophoresis. Detecting viral RNA or proviral DNA requires nucleic acid amplification tests like PCR, while reverse transcriptase activity assays are primarily used in research.

Question 2

A 9-year-old boy is evaluated for intellectual disability, a long narrow face, prominent jaw, and large ears. His family history is significant for a maternal uncle with similar features. To confirm the suspected diagnosis of Fragile X syndrome, a genetic test is performed. The technique involves digesting the patient's DNA with a restriction enzyme, separating the fragments by gel electrophoresis, and using a labeled DNA probe corresponding to the FMR1 gene.

Which of the following laboratory techniques was most likely used to detect the CGG trinucleotide repeat expansion in this patient?

  1. Southern blot (correct answer)
  2. Northern blot
  3. Western blot
  4. ELISA

Explanation: A Southern blot is used to detect specific sequences within a DNA sample. The key steps are restriction enzyme digestion of DNA, separation by size via gel electrophoresis, transfer to a membrane, and hybridization with a specific DNA probe. This method is classically used to identify the large CGG repeat expansions in the FMR1 gene characteristic of Fragile X syndrome. Northern blot analyzes RNA, Western blot analyzes protein, and ELISA detects antigens or antibodies.

Question 3

A 22-year-old sexually active woman presents to a public health clinic requesting screening for sexually transmitted infections. A rapid screening test for HIV is performed using a serum sample. The test device contains wells coated with recombinant HIV antigens. A colorimetric reaction occurs if the patient's sample contains the molecule of interest.

This screening test, known as an indirect ELISA, is designed to directly detect which of the following in the patient's serum?

  1. Host antibodies against HIV (correct answer)
  2. HIV p24 antigen
  3. HIV RNA
  4. CD4+ T-lymphocytes

Explanation: An indirect Enzyme-Linked Immunosorbent Assay (ELISA) for HIV screening works by detecting host-produced antibodies (IgG and IgM) against HIV. The test wells are coated with viral antigens. If the patient's serum contains anti-HIV antibodies, they will bind to these antigens. A secondary, enzyme-linked antibody that binds to human antibodies is then added to produce a detectable signal. Detection of p24 antigen uses a different type of ELISA (sandwich ELISA). HIV RNA is detected by RT-PCR. CD4+ cells are quantified by flow cytometry.

Question 4

A 3-week-old infant is brought to the emergency department with fever, poor feeding, and a high-pitched cry. On examination, vesicular lesions are noted on the scalp. A lumbar puncture is performed due to concern for neonatal herpes simplex virus (HSV) encephalitis. The cerebrospinal fluid is sent for the most sensitive and rapid diagnostic test.

Which of the following techniques, based on the amplification of nucleic acids, is the gold standard for diagnosing HSV infection in the CSF?

  1. Polymerase chain reaction (PCR) (correct answer)
  2. Viral culture
  3. Tzanck smear
  4. Serology for anti-HSV antibodies

Explanation: Polymerase chain reaction (PCR) is the diagnostic method of choice for HSV encephalitis. It is highly sensitive and specific, capable of detecting minute amounts of HSV DNA in the cerebrospinal fluid (CSF) within hours. Viral culture is too slow for this emergent condition. Tzanck smear lacks sensitivity and is used for skin lesions. Serology is not useful for diagnosing acute CNS infection as antibody presence does not distinguish between acute and past infection.

Question 5

A 40-year-old traveler returns from Southeast Asia with high fever, severe myalgia, and cough. A nasopharyngeal swab is sent for testing to diagnose influenza. Because influenza virus has an RNA genome, the laboratory must first synthesize a DNA strand using the viral genome as a template before proceeding with amplification.

The enzyme used in the initial step of this diagnostic assay is unique to which of the following techniques?

  1. Reverse transcriptase-PCR (RT-PCR) (correct answer)
  2. Standard PCR
  3. Southern blot
  4. Restriction fragment length polymorphism (RFLP)

Explanation: The detection of RNA viruses like influenza requires Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR). The key first step is the use of the enzyme reverse transcriptase to create a complementary DNA (cDNA) copy from the viral RNA template. This cDNA is then amplified using standard PCR methodology. Standard PCR, Southern blot, and RFLP all operate on DNA templates and do not involve reverse transcription.

Question 6

A neonate is born with a cleft palate, a ventricular septal defect, and hypocalcemic seizures. Physical examination reveals low-set ears and a small jaw. A geneticist suspects DiGeorge syndrome. A standard G-banded chromosome analysis is performed and appears normal. A more targeted test using a DNA probe labeled with a fluorescent dye that binds to chromosome 22q11.2 is ordered.

Which of the following techniques is most appropriate for detecting the microdeletion associated with this syndrome?

  1. Fluorescence in situ hybridization (FISH) (correct answer)
  2. Southern blot
  3. High-resolution karyotyping
  4. PCR

Explanation: Fluorescence in situ hybridization (FISH) is a molecular cytogenetic technique that uses fluorescent probes to detect specific DNA sequences on chromosomes. It is ideal for identifying submicroscopic deletions or duplications, such as the 22q11.2 deletion in DiGeorge syndrome, which are often too small to be seen on a standard or even high-resolution karyotype.

Question 7

A 40-year-old woman undergoes amniocentesis during her pregnancy due to advanced maternal age. In the laboratory, fetal cells from the amniotic fluid are cultured. A mitotic inhibitor is added to arrest the cells in metaphase. The cells are then harvested, spread on a slide, and stained with a specific dye that creates a pattern of light and dark bands on the chromosomes, which are then arranged by size and shape for analysis.

This entire process is used to generate which of the following?

  1. A karyotype (correct answer)
  2. A DNA fingerprint
  3. A gene expression profile
  4. A pedigree chart

Explanation: The described process of culturing cells, arresting them in metaphase, and staining and arranging the chromosomes is known as karyotyping. It provides a visual representation of the entire chromosome complement of an individual and is used to detect large-scale numerical (e.g., trisomy 21) or structural abnormalities.

Question 8

A 4-year-old boy presents with progressive muscle weakness. A muscle biopsy is obtained to analyze the dystrophin protein, which is absent in Duchenne muscular dystrophy. The protein extract from the biopsy is treated with an anionic detergent to denature the proteins and give them a uniform negative charge. The sample is then loaded onto a gel and subjected to an electric field.

The separation of proteins in this technique, known as SDS-PAGE, is based primarily on which property?

  1. Molecular weight (correct answer)
  2. Isoelectric point
  3. Native charge
  4. Subcellular localization

Explanation: In sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the detergent SDS denatures proteins and coats them with a uniform negative charge, eliminating the influence of their native charge. As a result, when an electric current is applied, the proteins migrate through the gel matrix solely based on their size (molecular weight), with smaller proteins moving faster and further.

Question 9

A 10-month-old infant of Mediterranean descent is evaluated for severe microcytic anemia. Hemoglobin electrophoresis is performed to investigate for beta-thalassemia. The test involves applying a sample of the patient's red blood cell lysate to a gel strip that contains a stable pH gradient. An electric field is applied, and different hemoglobin variants migrate until they reach the pH where their net charge is zero.

This method of separating proteins based on their pI is known as which of the following?

  1. Isoelectric focusing (correct answer)
  2. SDS-PAGE
  3. Size-exclusion chromatography
  4. Affinity chromatography

Explanation: Isoelectric focusing (IEF) is a technique that separates proteins based on their isoelectric point (pI). Proteins migrate through a pH gradient in an electric field until they reach the pH equal to their pI, at which point they have no net charge and stop moving. This is particularly useful for separating hemoglobin variants that may differ by a single amino acid, altering their overall charge.

Question 10

A research laboratory is developing a novel therapy based on a recombinant protein. The protein is expressed in a bacterial culture system with an engineered glutathione-S-transferase (GST) tag. To purify the protein, the cell lysate is passed through a column containing a resin with glutathione immobilized on its surface.

This purification technique, which exploits a specific binding interaction, is an example of which of the following?

  1. Affinity chromatography (correct answer)
  2. Ion-exchange chromatography
  3. Size-exclusion chromatography
  4. Gel filtration chromatography

Explanation: Affinity chromatography is a method of separating a specific molecule from a mixture based on a highly specific interaction, such as that between an enzyme and substrate, or an antigen and antibody. In this case, the GST-tagged protein specifically binds to the immobilized glutathione ligand on the column resin, while all other proteins wash through. The purified protein can then be eluted by adding free glutathione.

Question 11

A 48-year-old man with chronic hepatitis C is treated with a direct-acting antiviral regimen. To monitor the effectiveness of the treatment, his physician orders a test to measure the amount of viral genetic material in the blood. The test uses a fluorescent probe that binds to the amplified viral product, allowing for the measurement of its accumulation with each cycle of amplification.

This method for determining viral load is best described as which of the following?

  1. Quantitative PCR (qPCR) (correct answer)
  2. ELISA
  3. Western blot
  4. Sanger sequencing

Explanation: Quantitative PCR (qPCR), or real-time PCR, is used to quantify the amount of a specific nucleic acid in a sample. By monitoring the fluorescence signal generated during each amplification cycle, the initial quantity of the target sequence (e.g., viral RNA after reverse transcription) can be determined. This is the standard method for measuring viral load in infections like HCV and HIV. ELISA and Western blot detect proteins/antibodies, while Sanger sequencing determines nucleotide sequences.

Question 12

To study the function of a liver-specific enzyme in glucose metabolism, scientists create a conditional knockout mouse. They first create a mouse line in which the gene for the enzyme is flanked by loxP sequences. This mouse is then cross-bred with a mouse line that expresses Cre recombinase under the control of an albumin promoter, which is active only in hepatocytes.

In the liver cells of the resulting offspring, the Cre recombinase will catalyze which of the following molecular events?

  1. Excision of the gene flanked by loxP sites (correct answer)
  2. Activation of the gene flanked by loxP sites
  3. Methylation of the albumin promoter
  4. Insertion of a new gene at the loxP sites

Explanation: The Cre-Lox system is a tool for site-specific recombination. The Cre recombinase enzyme recognizes and cuts at specific DNA sequences called loxP sites. When a gene segment is flanked by two loxP sites oriented in the same direction, Cre recombinase will excise the intervening DNA, leading to a gene knockout specifically in the tissues where Cre is expressed (in this case, hepatocytes).

Question 13

Scientists are attempting to correct a single-base-pair mutation in a gene that causes a rare metabolic disorder. They design a system consisting of a nuclease protein and a synthetic RNA molecule. The RNA molecule contains a sequence that is complementary to the target DNA sequence near the mutation, guiding the nuclease to create a double-strand break at that specific site.

In this CRISPR-Cas9 gene-editing system, the target specificity is primarily determined by which component?

  1. The guide RNA sequence (correct answer)
  2. The Cas9 protein's enzymatic domain
  3. The protospacer adjacent motif (PAM) sequence
  4. The cell's DNA repair machinery

Explanation: The specificity of the CRISPR-Cas9 system—its ability to target a precise location in the genome—is determined by the nucleotide sequence of the guide RNA (gRNA). The gRNA forms Watson-Crick base pairs with the target DNA strand, directing the Cas9 nuclease to that location. While the PAM sequence is necessary for Cas9 to bind and cut, the gRNA provides the specific address.

Question 14

A newborn screening test for sickle cell anemia is performed. DNA is extracted from a blood spot, and the region of the beta-globin gene containing the mutation is amplified. This amplified DNA is then incubated with the restriction enzyme DdeI. The normal allele has two DdeI cutting sites, while the sickle cell allele has only one because the point mutation eliminates a site. The resulting fragments are analyzed by gel electrophoresis.

This diagnostic approach, which relies on a mutation affecting an enzyme recognition site, is an example of which technique?

  1. Restriction fragment length polymorphism (RFLP) (correct answer)
  2. Allele-specific oligonucleotide (ASO) hybridization
  3. Sanger sequencing
  4. Karyotyping

Explanation: Restriction fragment length polymorphism (RFLP) is a technique that exploits variations in homologous DNA sequences, known as polymorphisms, that affect restriction enzyme cleavage sites. The single nucleotide polymorphism causing sickle cell disease abolishes a restriction site, leading to a different pattern of DNA fragments after enzyme digestion, which can be visualized on a gel. This difference in fragment length allows for diagnosis.

Question 15

A researcher is purifying a mixture of proteins at a buffered pH of 7.4. The mixture contains albumin (pI = 4.7) and myoglobin (pI = 7.0). The researcher uses a chromatography column containing a positively charged diethylaminoethyl (DEAE) cellulose resin.

Under these conditions, which of the following is the most likely outcome?

  1. Albumin will bind more strongly to the column than myoglobin (correct answer)
  2. Myoglobin will bind more strongly to the column than albumin
  3. Both proteins will bind equally to the column
  4. Neither protein will bind to the column

Explanation: This is an example of anion-exchange chromatography, where a positively charged resin (DEAE) binds negatively charged molecules. At pH 7.4, both proteins will have a net negative charge since the pH is above their respective pIs. However, albumin (pI = 4.7) will be much more negatively charged than myoglobin (pI = 7.0) at this pH. The further a protein's pI is from the buffer pH, the greater its net charge. Therefore, albumin will bind much more strongly to the positively charged column.

Question 16

A 5-year-old boy presents with pallor, easy bruising, and recurrent fever. A complete blood count shows pancytopenia with circulating blasts. A bone marrow aspirate is performed, and the sample is treated with fluorescently-labeled antibodies against various cell surface markers. The cells are then passed one-by-one through a laser beam to identify their lineage and maturation stage, which is crucial for diagnosis and classification of acute leukemia.

Which of the following laboratory techniques is described in this scenario?

  1. Flow cytometry (correct answer)
  2. Immunohistochemistry
  3. ELISA
  4. Karyotyping

Explanation: Flow cytometry is a technique used to analyze the characteristics of individual cells within a heterogeneous population. By using fluorescent antibodies against specific surface proteins (CD markers), it can identify and quantify cell populations, which is essential for the diagnosis and classification of hematologic malignancies like acute lymphoblastic leukemia (ALL) or acute myeloid leukemia (AML).

Question 17

A pathologist is studying a breast tumor biopsy to determine its molecular subtype, which has important prognostic and therapeutic implications. The pathologist wants to compare the expression levels of thousands of genes, including estrogen receptor, progesterone receptor, and HER2/neu, simultaneously between the tumor tissue and normal adjacent tissue. cDNAs from both samples are fluorescently labeled and applied to a glass slide containing thousands of known DNA spots.

Which of the following technologies is best suited for this comprehensive gene expression analysis?

  1. DNA microarray (correct answer)
  2. Northern blot
  3. In situ hybridization
  4. Quantitative PCR

Explanation: DNA microarray technology allows for the simultaneous measurement of the expression levels of thousands of genes. This is ideal for creating a global gene expression profile of a tumor, which can be used for classification and prognosis. While Northern blot and qPCR can measure gene expression, they are typically used for one or a few genes at a time. In situ hybridization shows location but is less quantitative for a large number of genes.

Question 18

Researchers are investigating the mechanism of a new chemotherapy agent on a specific type of lung cancer cell. They hypothesize that the drug works by downregulating the transcription of an oncogene essential for cell proliferation. To test this, they treat the cells with the drug, isolate total cellular RNA, and then use a technique to separate the RNA by size and detect the specific mRNA transcript of interest with a labeled probe.

Which of the following techniques is most appropriate for assessing the abundance of the specific mRNA transcript in this experiment?

  1. Northern blot (correct answer)
  2. Southern blot
  3. Microarray
  4. Sanger sequencing

Explanation: A Northern blot is a laboratory technique used to detect and quantify specific RNA molecules (such as mRNA) in a sample. It directly measures the amount and size of the transcript of interest. While a microarray can also measure mRNA levels, it assesses thousands of genes simultaneously and is less focused than a Northern blot for a single target. Southern blot analyzes DNA, and Sanger sequencing determines the nucleotide sequence.

Question 19

A biochemist aims to isolate a large protein complex with a molecular weight of 500 kDa from a mixture of smaller proteins, including monomeric enzymes (~50 kDa) and peptides (~5 kDa). The mixture is applied to a long column packed with porous beads. The buffer flows through the column, and fractions are collected as they exit.

Using this size-exclusion chromatography technique, in which fractions would the 500 kDa protein complex be expected to appear?

  1. In the first fractions to elute (early fractions) (correct answer)
  2. In the last fractions to elute (late fractions)
  3. In the middle fractions, after the peptides but before the enzymes
  4. Spread evenly across all fractions

Explanation: In size-exclusion chromatography (also known as gel filtration), large molecules are excluded from the pores of the chromatography beads and thus travel through the column in the volume outside the beads (the void volume). This is a shorter path, so they elute first. Smaller molecules can enter the pores, increasing their effective path length and causing them to elute later. Therefore, the largest molecule, the 500 kDa complex, will elute in the earliest fractions.

Question 20

A newborn has a positive screening test for cystic fibrosis. A subsequent sweat chloride test is equivocal. To confirm the diagnosis and identify the specific genetic lesion, a blood sample is sent for analysis of the CFTR gene. The test utilizes dideoxynucleotides labeled with different fluorescent dyes to terminate DNA synthesis at each base, allowing for the determination of the exact base-by-base sequence of the gene.

Which of the following techniques was used to identify the precise mutation in this infant's CFTR gene?

  1. Sanger sequencing (correct answer)
  2. Karyotyping
  3. Western blot
  4. Southern blot

Explanation: Sanger sequencing is the method described. It determines the precise order of nucleotides in a DNA molecule by using chain-terminating dideoxynucleotides. This allows for the identification of specific point mutations, deletions, or insertions within a gene, such as the common ΔF508 mutation in cystic fibrosis. Karyotyping assesses whole chromosomes, Western blot detects proteins, and Southern blot detects specific DNA sequences without providing base-by-base resolution.