IB Biology HLPractical SkillsPaper 1B & IA~16 min read
Applying Lab Techniques
There are nine techniques the course expects you to know, and the requirement is always the same two things: describe the purpose and explain how to carry it out. Nobody is asking you to memorise a method word for word — they are asking whether you understand what the technique is for.
📚 What you need to know
Paper or thin layer chromatography — separating a mixture, and calculating Rf values.
Colorimetry and serial dilutions — measuring concentration from absorbance, using a calibration curve.
Physical and digital molecular modelling — representing molecules in three dimensions.
Light microscope and eyepiece graticule — measuring specimens after calibration against a stage micrometer.
Preparation of temporary mounts — slides made to view living or fresh material.
Identifying and classifying organisms — using dichotomous keys.
Sampling techniques — random versus systematic sampling, with quadrats and transects.
Karyotyping and karyograms — arranging chromosomes to detect abnormalities.
Cladogram analysis — reading evolutionary relationships from branching diagrams.
1. Chromatography
Purpose: to separate the components of a mixture — classically the photosynthetic pigments in a leaf — and to identify them.
Separation happens because each component has a different solubility in the solvent and a different attraction to the paper. The more soluble a component is, and the less it is held by the paper, the further it travels.
🧩 Carrying it out
Draw a baseline in pencil near the bottom of the paper. Pencil is used because ink would dissolve and separate too.
Apply a small, concentrated spot of the mixture to the baseline, letting it dry between applications.
Stand the paper in a small volume of solvent, making sure the solvent level is below the baseline, or the spot will simply dissolve into it.
Cover the container so the atmosphere becomes saturated with solvent vapour and the solvent does not evaporate from the paper.
Remove the paper before the solvent front reaches the top, and mark the front immediately in pencil.
Measure from the baseline to the centre of each spot, and to the solvent front.
Retardation factor
Rf = distance moved by the spot ÷ distance moved by the solvent front
Rf has no units, because it is a distance divided by a distance, and it is always less than 1. For a given solvent and paper it is a constant for each substance, so it can be used to identify components by comparing with known values.
If your solvent runs off the top of the paper you cannot calculate anything, because there is no measurable solvent front. That single mistake ruins more chromatography practicals than any other.
2. Colorimetry and serial dilutions
Purpose: to measure the concentration of a coloured solution, or how much light a suspension blocks, by measuring how much light passes through it.
A colorimeter shines light of a chosen wavelength through a sample in a cuvette and measures the absorbance (or transmission). The more concentrated the coloured solution, the more light is absorbed. The instrument must first be zeroed with a blank — a cuvette containing only the solvent.
A colorimeter reading on its own means nothing. To convert absorbance into concentration you need a calibration curve, and to build one you need a set of solutions of known concentration — which is what a serial dilution gives you.
Serial dilution is used because it is far more accurate than trying to measure out a tiny volume directly. Making 0.0001 mol dm−3 in one step would mean measuring 0.0025 cm3 of stock, which no school apparatus can do.
Once you have the dilutions, measure the absorbance of each, plot absorbance against known concentration, and draw a line of best fit. That is your calibration curve. An unknown sample can then be read off it: find its absorbance on the y-axis, go across to the line, and read the concentration below.
3. Physical and digital molecular modelling
Purpose: to represent molecules in three dimensions so that shape, bonding and interactions can be understood.
Physical models — ball-and-stick kits — make bond angles and the arrangement of atoms tangible. Digital models can be rotated, zoomed and stripped back to show only a backbone or an active site, and can handle molecules far too large to build by hand, such as a whole enzyme.
The evaluation point examiners like: every model is a simplification. A ball-and-stick model shows connectivity clearly but wildly exaggerates the empty space in a molecule; a space-filling model shows volume correctly but hides the bonds.
4. The light microscope and eyepiece graticule
Purpose: to measure the actual size of a specimen seen down a microscope.
An eyepiece graticule is a tiny scale inside the eyepiece. It stays the same apparent size whatever objective lens you use, so its divisions have no fixed value — the same division covers a different real distance at each magnification. That is why it has to be calibrated first, against a stage micrometer: a slide with a scale of known size etched onto it, usually with divisions of 10 µm.
The stage micrometer is only needed once per magnification. After that it comes off the stage and the graticule does all the work.
5. Preparation of temporary mounts
Purpose: to view fresh or living material under a microscope without permanently preserving it.
Cut or peel a very thin section — thin enough for light to pass through.
Place it in a drop of water or stain on a clean slide.
Lower a coverslip at an angle, using a mounted needle, so air bubbles are pushed out ahead of it. Bubbles look like thick dark circles and are often mistaken for cells.
Add stain at one edge and draw it through with filter paper on the opposite edge if staining after mounting.
Common stains include iodine for starch, and methylene blue for animal cell nuclei. Stains increase contrast, because most cell components are transparent.
6. Identifying and classifying organisms
Purpose: to identify an unknown organism from its observable features.
The standard tool is a dichotomous key — a series of paired statements, each pair splitting the remaining possibilities in two. Start at the first pair, choose the statement that matches your specimen, and follow it to the next pair or to a name.
Good keys use features that are constant and easy to observe — number of legs, leaf shape, presence of a shell. Poor keys use features that vary with age, season or individual, such as overall size or colour intensity.
7. Sampling techniques
Purpose: to estimate the abundance or distribution of organisms without counting every individual.
Random sampling avoids bias so the sample represents the whole area. Systematic sampling deliberately introduces a pattern, because the pattern is the thing being investigated — how a community changes from the low shore to the high shore, for example.
Random sampling uses random number pairs as coordinates on a grid laid over the area. It avoids bias and is used when the habitat appears uniform.
Systematic sampling places quadrats at regular intervals, usually along a transect line. It is used when there is an environmental gradient — distance up a shore, distance from a path, light intensity under a canopy.
Abundance is recorded as a count, or as percentage cover for organisms that cannot be counted individually, such as grasses or lichens.
8. Karyotyping and karyograms
Purpose: to examine an individual’s chromosomes for number and structure.
Cells are arrested in metaphase, when chromosomes are most condensed and visible, then stained and photographed. A karyogram is the resulting image with the chromosomes cut out and arranged in homologous pairs, ordered by size and by the position of the centromere.
From a karyogram you can determine biological sex (from the sex chromosomes) and detect abnormalities in chromosome number, such as a trisomy where three copies of one chromosome are present instead of two.
9. Cladogram analysis
Purpose: to show and interpret the evolutionary relationships between groups of organisms.
A cladogram is a branching diagram built from shared derived characteristics, increasingly from base sequence or amino acid sequence data. Each node is a common ancestor, and each branch point represents a divergence.
The key skill is reading relatedness correctly: two groups are more closely related if they share a more recent common ancestor. Position along the top of the diagram means nothing — branches can be rotated at any node without changing what the cladogram says.
Technique
Its purpose
The detail examiners look for
Chromatography
Separate and identify components of a mixture
Pencil baseline, solvent below the line, Rf to the spot centre
Colorimetry
Find concentration from absorbance
Zero with a blank, and use a calibration curve
Serial dilution
Make a range of known concentrations
The dilution factor at each step, and mixing between steps
Molecular modelling
Represent molecules in three dimensions
What the model simplifies or exaggerates
Eyepiece graticule
Measure specimen size down a microscope
Calibration against a stage micrometer at each magnification
Temporary mounts
View fresh material under a microscope
Thin section, coverslip lowered at an angle, stain for contrast
Dichotomous keys
Identify an unknown organism
Paired statements based on constant, observable features
Sampling
Estimate abundance and distribution
Random for uniform areas, systematic for gradients
Karyograms
Examine chromosome number and structure
Metaphase cells, arranged by size and centromere position
Cladograms
Show evolutionary relationships
Relatedness read from the most recent common ancestor
Read the command term before you start writing. Describe the purpose wants one or two sentences on what the technique is for and why it exists. Explain how to carry it out wants the steps, with the reason for the fiddly ones — why the baseline is pencil, why the container is covered, why you calibrate again after changing lens.
Worked examples
WE 1
Calculating and using an Rf value
A leaf pigment separates into a spot 6.3 cm from the baseline; the solvent front is 9.0 cm from the baseline. In this solvent, known Rf values are: carotene 0.95, xanthophyll 0.70, chlorophyll a 0.65, chlorophyll b 0.45. Calculate the Rf and identify the pigment. (3 marks)
Step 1: substituteRf = 6.3 ÷ 9.0Step 2: calculateRf = 0.70 (no units)Step 3: identify
This matches the value for xanthophyll.
Step 4: the necessary caution
Rf values are only comparable if the same solvent and paper were used, so the identification depends on the reference values coming from identical conditions.
Rf = 0.70, so the pigment is xanthophyllRf never has units and is never greater than 1. If you get 1.4, you have divided the wrong way round.
WE 2
Calibrating a graticule
Under the ×40 objective, 100 eyepiece graticule divisions line up exactly with 40 divisions of a stage micrometer. Each stage micrometer division is 10 µm. A cell measures 26 eyepiece divisions across. Calculate its actual width. (4 marks)
Step 1: find the real distance covered40 × 10 = 400 µmStep 2: find the value of one eyepiece division400 ÷ 100 = 4 µm per divisionStep 3: measure the cell26 × 4 = 104 µmStep 4: the point of the exercise
If the objective is changed, the eyepiece divisions stay the same apparent size but now cover a different real distance, so the calibration must be repeated.
The cell is 104 µm widecheck your answer is sensible. Most plant cells are tens to hundreds of micrometres across, so 104 µm is believable; 104 mm would not be.
WE 3
Planning a dilution
A student has a 2.0 mol dm−3 stock solution and needs 25 cm3 of a 0.4 mol dm−3 solution. Calculate the volume of stock required, and state how to prepare it. (3 marks)
Step 1: use the dilution relationshipc1V1 = c2V2, so V1 = (0.4 × 25) ÷ 2.0Step 2: calculateV1 = 10 ÷ 2.0 = 5.0 cm3 of stockStep 3: state the method
Measure 5.0 cm3 of stock with a pipette, add 20.0 cm3 of distilled water to make the total up to 25 cm3, and mix thoroughly.
Step 4: the check
The solution has been diluted five-fold, and 2.0 ÷ 5 = 0.4, which confirms the answer.
5.0 cm3 of stock plus 20.0 cm3 of waterthe classic error is adding 25 cm3 of water to the stock, giving 30 cm3 of the wrong concentration. Make the volume up to the target, do not add the target.
💡 Exam tips
For every technique, be ready to state its purpose in one sentence before describing any steps.
Learn the reasons behind the fiddly details — pencil baseline, solvent below the line, covered container, blank in the colorimeter. Those are where the marks sit.
Rf is measured to the centre of the spot, has no units, and is always less than 1.
Graticule calibration must be redone at every magnification.
Justify the sampling method by the habitat: uniform means random, gradient means systematic.
On a cladogram, relatedness comes from the most recent common ancestor, never from how close two labels appear along the top.
⚠ Common mistakes
Drawing the chromatography baseline in pen. The ink separates and ruins the run.
Starting the solvent above the baseline. The spot dissolves into the solvent instead of travelling up the paper.
Measuring Rf to the top or edge of a spot. Always the centre.
Forgetting to zero the colorimeter with a blank. Every reading is then offset by the same amount — a systematic error.
Assuming a graticule division is a fixed length. It only has meaning once calibrated, at that magnification.
Choosing quadrat spots “randomly” by eye. That is not random. Use random number coordinates.
Reading a cladogram left to right. Branches can be rotated freely; only the nodes carry information.
Pulling the skill set together
Three pages, one question repeated in different forms: can this procedure be trusted? Safety and ethics decide whether it should be done at all. Measurement decides whether the numbers mean anything. Technique decides whether the method actually answers the question you asked.
Where these marks appear. Not just in Paper 1B. Every one of these ideas is assessed again in your internal assessment, where “explain how you minimised risk” and “evaluate the precision of your measurements” are worth real credit — and where a generic answer stands out immediately.
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