Domain: Healthcare — Basic Immunology
Target concept: B Lymphocyte Detection (Surface Antibody Staining)

Video generation prompt: B lymphocytes are detected by surface antibody staining. The procedure involves adding labeled antibodies to a prepared cell suspension, gently mixing, incubating, washing, and then observing under a fluorescence microscope or flow cytometer.

Evaluation guide:
(i) Target concept: Specific detection of B lymphocytes via surface antigen binding of fluorophore-labeled antibodies, requiring antigen-antibody binding, unbound antibody removal, and fluorescence visualization.
Minimum mechanism: Antibody must be physically present and contact cells; binding requires time (incubation); non-specific antibody must be removed (washing); fluorescence excitation/emission must be isolated from background (achieved by washing + instrument optics).
(ii)Storyline: Frame 1–2: Labeled antibody vial and cell suspension tube appear side-by-side in fixed view. Frame 3–4: Pipette draws from vial and dispenses into tube (contact confirmed). Frame 5–6: Tube undergoes gentle circular swirling (continuous motion, no spill). Frame 7: Tube remains motionless (stillness confirms incubation pause). Frame 8: Pipette performs aspiration/ejection cycle (wash motion). Frame 9–10: Microscope/flow cytometer screen activates, displaying fluorescent cells or a shifted population histogram — signal appears only now, localized to discrete cellular objects, with no pre-existing fluorescence in earlier frames. All events occur from fixed frontal viewpoint; no object leaves frame; lighting remains constant; no zoom/pan.

Manual:
Experimental Procedure
1. Add fluorophore-labeled anti-B-cell antibody to cell suspension tube.
2. Gently mix by swirling.
3. Incubate in dark (≥15 min).
4. Wash cells by centrifugation and buffer resuspension.
5. Observe under fluorescence microscope or flow cytometer.
Observation of Experimental Phenomena
Fluorescent signal appears exclusively on intact, round B lymphocytes — no signal on debris or background; signal is bright, punctate, and membrane-localized.
Summary of Experimental Principles
B lymphocytes express surface immunoglobulins (e.g., CD19, CD20); fluorophore-conjugated antibodies bind specifically to these markers; after washing, only bound antibody remains, enabling detection via fluorescence.

Prompt Grounding rubric:
level_1: No labeled antibody vial, no cell suspension tube, no pipette, no wash/centrifuge step, and no fluorescence microscope or flow cytometer screen appear anywhere; none of the five procedural actions -- antibody addition, mixing, incubation, washing, observation -- is depicted or recognizable.
level_2: At least one core object appears (e.g., the tube or the vial) but with none of the five actions correctly attached to it; or two objects appear (e.g., tube and pipette) with an incorrect spatial relationship, such as the pipette never actually contacting the tube; no coherent step-by-step sequence of antibody addition, mixing, incubation, washing, or observation can be identified.
level_3: All five core objects are present and identifiable: the labeled antibody vial, the cell suspension tube, the pipette, a visible wash step (e.g., buffer exchange or a spin-down implied by tube handling), and a fluorescence microscope or flow cytometer display; at least three of the five actions -- (1) pipette dispensing antibody into the tube, (2) swirling/mixing, (3) a static incubation pause, (4) a washing action, (5) a final screen showing a readout -- appear in the correct order, with at most one action missing or out of place.
level_4: All five core objects are present and correctly identifiable by shape and function -- the vial as the antibody source, the tube holding the cell suspension, the pipette used for transfer, a visible wash step, and a microscope/cytometer screen for the readout; all five actions occur in the correct order with none omitted, duplicated, or reordered; the tube and the screen each stay in a consistent on-screen location across the sequence; only non-core flaws are allowed, such as unreadable text on the vial's label or a screen background that doesn't look distinctly fluorescent.
level_5: Every object is unambiguous and identifiable by both form and function -- the vial is visibly labeled or color-coded for a specific fluorophore, the tube shows a visible cell pellet before the wash step, the pipette tip is clearly seen entering and leaving the tube, and the microscope/cytometer screen shows a clearly separated bright population (a histogram peak or dot-plot cluster) distinct from a dim or negative background; all five actions occur in strict, correctly ordered sequence with clear causal linkage -- mixing directly follows antibody addition, washing directly follows incubation; every label, object edge, and the on-screen readout are clearly legible and easy to make out at normal viewing size.

Scientific and Causal Correctness rubric:
level_1: No fluorescent signal appears at the final observation step; the cell suspension looks uniformly unstained the whole time; the antibody vial shows no color cue tied to a fluorophore (e.g., it looks clear or has an ordinary, non-glowing tint); there's no apparent link between adding the antibody and any signal appearing later.
level_2: The final observation shows scattered bright specks or generic noise rather than signal confined to individual, cell-sized objects; the antibody vial's color doesn't match any real fluorophore (e.g., a red liquid in a vial that's supposed to be FITC-labeled); mixing happens but nothing follows from it; an incubation pause is shown but with no meaningful time gap before observation; or the wash step is skipped even though a hazy background glow appears in the final image.
level_3: The antibody vial's color is consistent with a real fluorophore (green for FITC, orange for PE, or deep red for APC); the tube looks uniformly opaque before staining, and after mixing, incubation, and washing, the final observation shows fluorescent signal confined to discrete, cell-sized objects rather than a diffuse haze; the signal only appears after the wash step, not before it; the chain of addition -> mixing -> incubation -> washing -> visible signal on cells is mostly intact, with at most one break, such as signal showing up before washing is complete.
level_4: The antibody vial's color matches a standard fluorophore convention (green for FITC, orange-red for PE, or far-red for APC); the tube visibly moves through recognizable states -- an opaque suspension, brief turbidity while mixing, a settled pellet after the wash step, and a clear liquid above it; the final observation shows discrete, fluorescent, cell-shaped objects of consistent size with clear edges; signal only ever appears after the full addition -> mix -> incubate -> wash -> observe sequence, with no signal in the tube before antibody addition and no background glow on the screen before observation; only non-core flaws are allowed, such as slightly oversaturated color or minor color bleed.
level_5: The vial's color is unmistakably tied to one specific fluorophore (a bright apple-green for FITC or a vivid orange for PE) and could plausibly carry a label like 'anti-CD19-FITC'; the tube visibly passes through a translucent milky suspension, a uniformly turbid swirl while mixing, gentle pelleting after incubation, and a clear liquid with a compact pellet after washing; the final readout shows a high-contrast fluorescent population -- a clearly right-shifted peak on a cytometer histogram, or bright round cells against a dim background on a microscope screen; every state change and the antibody-to-signal causal chain is easy to follow and matches how real immunostaining behaves, with no unexplained mismatch in color, timing, or where the signal shows up.

Spatiotemporal Consistency rubric:
level_1: Objects behave impossibly between frames: the pipette appears already inside the tube with no visible entry motion, the cell suspension tube is present and then is simply gone before reappearing with no motion connecting the two states, the microscope/cytometer screen's content flickers inconsistently, the overall lighting suddenly jumps between dark and bright, or the tube or pipette visibly breaks or bends in an unnatural way.
level_2: At least one clear discontinuity appears: the pipette tip enters the tube off-frame and reappears already inside it; the tube rotates a large angle with no in-between motion shown; the screen's content changes with no visible transition, such as jumping straight from blank to a full histogram; the lighting shifts noticeably between steps; or the motion visibly skips or stutters more than once.
level_3: The tube, pipette, vial, and screen stay in believable positions relative to each other throughout -- for instance the tube stays centered, the pipette stays beside it, the screen stays in the same corner of the frame; motion within each action looks continuous, with the pipette lowering smoothly and the swirl tracing an unbroken circular path; the room lighting on the tube and pipette doesn't shift noticeably between steps; no object looks stretched, warped, or broken; transitions between steps use visible motion rather than an abrupt cut. One small flaw is allowed, such as a brief stutter during the swirling motion or a moment of soft focus during the wash step.
level_4: No teleporting objects, no lighting jumps, and no visible structural distortion appear anywhere; all motion looks physically natural -- the swirl behaves like a real fluid and the pipette speeds up and slows down the way a real hand-held pipette would; the camera viewpoint stays fixed with no pan, tilt, or zoom; every object keeps a consistent size, orientation, and layering relative to the others throughout. Only very minor, non-core flaws are allowed, such as a faint brightening or dimming a viewer would barely notice, or a light trailing smear behind fast motion.
level_5: Spatiotemporal continuity looks essentially perfect to the eye: the pipette's path into and out of the tube is fully visible with no gaps; the swirling motion traces a smooth, even loop with no sudden speed changes; during the incubation pause the tube looks completely still, with no visible drift; the wash step shows a clear, coordinated aspirate-then-eject motion with the liquid surface moving continuously rather than jumping; the screen's content builds up gradually -- the histogram fills in or cells accumulate over time -- rather than swapping instantly; motion, lighting, and the tube and pipette's sense of depth and perspective all stay coherent and consistent across the whole sequence.