Biology 9700/13 — October/November 2019

Background Concept

A simple light microscope is limited in two ways: by magnification (how much bigger an image appears) and by resolution (the smallest distance between two points at which they can still be seen as separate). The resolution of a good light microscope with visible light is about 200 nm (0.2 µm). Anything smaller than this cannot be distinguished, no matter how much the image is enlarged — this is why electron microscopes (resolution ~0.2 nm) are needed for very small structures.

Common plant-cell structures and their typical sizes:

• Nucleus: ~5–20 µm — easily seen.
• Nucleolus: ~1–7 µm, a dense, darkly staining body inside the nucleus — clearly seen.
• Chloroplasts: ~3–10 µm long — visible as green ovals, but the internal grana (stacks of thylakoids, each ~0.3–0.6 µm) are too small to resolve as individual stacks at ×400.
• Lysosomes: ~0.1–1.2 µm — usually not visible without special staining.
• Ribosomes: ~20–30 nm — far below the resolution of a light microscope; need an electron microscope.

Staining (e.g. with methylene blue, iodine or acetocarmine) increases contrast by binding to particular cellular components, making them appear darker. The nucleolus, being rich in RNA and protein, takes up stain strongly and appears as a dark spot within the lighter nucleus.

Understanding the Question

The question specifies a simple light microscope (no phase contrast, no fluorescence, no electron microscopy) using daylight (white light) and a total magnification of ×400. We are asked which structure among the four options would be clearly visible under these conditions after staining. We must pick the structure that is both large enough to be resolved by a light microscope and that becomes obvious after routine staining.

Approach

Compare each option against the resolution limit of the light microscope and against the ability of routine stains to reveal it:

1. Is the structure larger than ~200 nm?
2. Does routine staining make it stand out clearly at ×400?

If the answer to both is yes, the option is correct.

Step-by-Step Reasoning

• A — chloroplast grana: Although whole chloroplasts are easily seen as green ovals, the grana (stacks of thylakoid membranes) are sub-microscopic at ×400. They cannot be individually resolved with a light microscope. Reject.
• B — lysosomes: Even when present (lysosomes are more typical of animal cells), they are small membrane-bound vesicles, often <1 µm, and do not stain distinctively with routine stains. Not clearly visible. Reject.
• C — nucleoli: Dense, RNA-rich bodies 1–7 µm across, sitting inside the nucleus. They take up basic dyes strongly and appear as dark, well-defined spots within a paler nucleus at ×400. Clearly visible. Correct.
• D — ribosomes: At ~20–30 nm, ribosomes are more than ten times smaller than the resolution limit of a light microscope. They cannot be seen at any magnification using visible light. Reject.

Key Takeaways

• The resolution of a light microscope (~200 nm) sets the lower size limit of what can be seen.
• Whole chloroplasts, nuclei and nucleoli are visible at ×400; grana, ribosomes and individual membrane proteins are not.
• Routine staining (e.g. methylene blue, acetocarmine) makes the nucleolus a particularly prominent feature of stained plant cells.

Common Mistakes

• Confusing whole chloroplasts (visible) with grana (not resolved) and assuming both can be seen.
• Choosing ribosomes because they are 'common in cells' — they are far too small for light microscopy.
• Assuming lysosomes can be seen simply because they exist; they are small and require special stains.

Things to Be Careful About

• The question asks what is clearly visible at ×400 with a simple light microscope and daylight — there is no phase contrast, fluorescence or electron microscopy available.
• Resolution, not magnification, is the limiting factor: increasing magnification without improving resolution gives only empty magnification (a larger blur, not more detail).