USMLE Step 1 Quiz: Biochemical Laboratory Techniques

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?

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?